Bangladesh Journal of

Pharmacology
Volume: 14; Number 4; Year 2019

Cite this article as: Kim DD, Nguyet VT, Anh HX, Trang NTT, Chuyen NH, Huong LM, Ha TTH, Ho DH, Dat
NT. Cytotoxic phenolic constituents from the leaves of Ehretia asperula. Bangladesh J Pharmacol. 2019; 14: 196-
97.
A Journal of the Bangladesh Pharmacological Society (BDPS) Bangladesh J Pharmacol 2019; 14: 196-197
Journal homepage: www.banglajol.info
Abstracted/indexed in Academic Search Complete, Agroforestry Abstracts, Asia Journals Online, Bangladesh Journals Online, Biological Abstracts,
BIOSIS Previews, CAB Abstracts, Current Abstracts, Directory of Open Access Journals, EMBASE/Excerpta Medica, Global Health, Google Scholar,
HINARI (WHO), International Pharmaceutical Abstracts, Open J-gate, Science Citation Index Expanded, SCOPUS and Social Sciences Citation Index
ISSN: 1991-0088; DOI: 10.3329/bjp.v14i4.42414

Letter to the Editor

Cytotoxic phenolic constituents from the determined by the HR-ESI-MS molecular ion peak at m/
leaves of Ehretia asperula z 259.0975 [M+H]+. The 1H NMR spectrum of 1 showed
an aromatic ABX spin system at δH 7.04 (1H, d, J = 8.0
Hz, H-5), 7.32 (1H, d, J = 2.0 Hz, H-2) and 7.34 (1H, dd, J
Sir, = 8.0, 2.0 Hz, H-6), an aromatic A2B2 protons at δH 7.15
The genus Ehretia is mainly distributes in tropical areas (2H, d, J = 8.0 Hz, H-3′, 5′) and 6.84 (2H, d, J = 8.0 Hz, H
of Asia, Africa, Northern America and exhibited -2′, 6′). In addition, an aldehyde proton at δH 9.71 (1H, s,
valuable pharmacologial properties (Li et al., 2010; H-7) and two methylene signals at δH 2.73 (2H, t, J = 7.0
Shukla and Kaur, 2018). In Vietnam, Ehretia asperula Hz, H-7′), 3.71 (2H, t, J = 7.0 Hz, H-8′) were also
Zoll. & Mort. has been used in traditional medicine for recognized. The 13C-NMR combined with the HSQC
the treatment of ulcer, tumors, liver disease and experiments indicated the presence of two aromatic
inflammation (Nguyen et al., 2017). To date, there are rings [δC 131.1 (C, C-1), 118.3 (CH, C-2), 147.2 (C, C-3),
few reports about the biological activities and chemical 151.8 (C, C-4), 116.1 (CH, C-5), 128.8 (CH, C-6), 158.5 (C,
composition of this plant. C-1′), 119.5 (CH, C-2′, 6′), 131.0 (CH, C-3′, 5′), 133.7 (C, C
-4′)], an aldehyde group [δC 193.1 (CH, C-7)] and a 2-
In the present study, the leaves of E. asperula were subsituted ethanol moiety [δC 39.4 (CH2, C-7′), 64.0
collected from the Bai Rong Hamlet, Thuong Tien
(CH2, C-8′)]. The HMBC correlations showed the
Commune, Kim Boi District, Hoa Binh Province. The air-
coupling from H-7 to C-1, C-2 and C-6, from H-2 and H
dried and powdered material (5.0 kg) was extracted in
methanol at room temperature for 3 days and repeated 3 -6 to C-7 suggesting the aldehyde group attached to C-1
times. The combined extracts were evaporated under reduced of the ABX aromatic ring. These data were similar to
pressure to obtain a crude residue (320 g), which was then re- those of 3,4-dihydroxybenzadehyde (protocatechuic
dissolved in water (2 L) and successively extracted with n- aldehyde) moiety (Ooike et al., 1997). The methylene
hexane and ethyl acetate, respectively. The organic layers protons H-7′ coupled to C-4′, C-3′ and C-5′ indicated the
were dried in vaccuo to give n-hexane and 34.4 g ethyl ace- ethanol moiety attached to C-3′ of the A2B2 aromatic
tate residues, respectively. Ethyl acetate was chromato- ring, which suggested the structure of 4-(2-hydroxyethyl)
graphed on a silica gel column eluted with solvent mixtures -phenol (tyrosol) moiety (Hussain et al., 2014). Based on
of dichloromethane and methanol (100/1, 50/1, 10/1, 5/1, 1/1 these data, the structure of 1 consisted of protocatechuic
and 1/100, v/v) to give 6 fractions F1-F6, respectively. The aldehyde and tyrosol fragments. The NOESY spectrum
fraction F2 was subjected to a silica gel column eluted with showed the cross peak between H-2 and H-2′, 6′ confirming
dichloromethane-methanol (5/1) to afford two subfractions that two fragments linked via C-3 and C-1' positions (Figure
F2.1 and F2.2). Subfraction F2.1 was separated on a silica gel 1). Thus, compound 1 was newly elucidated to be 4-hydroxy-
column, eluting with n-hexane-dichloromethane-methanol 3-[4-(2-hydroxyethyl)-phenoxy]-benzaldehyde.
(5/15/1, v/v/v) to obtain compound 1 (7.1 mg) and caffeic
acid (2) (37.5 mg). Methyl caffeate (3) (5.0 mg) was purified Cytotoxicity of the isolated compounds was evaluated
from subfraction F2.2 by a C18 column using methanol-water against human lung (Hep-G2, LU-1), cervical (HeLa),
(1/2, v/v). Fraction F3 was subjected to fractionation over breast (MCF-7), and rhabdomyosarcoma (RD) cancer
silica gel column, eluted with dichloromethane-acetone (10/1, cell lines (Table I). Compound 1 exhibited the strongest
v/v) to give three subfractions F3.1-F3.3. Subfraction F3.1 effect to Hep-G2, LU-1, HeLa and RD cells with IC50
was separated using silica gel column and further purified values in the range of 7.1-10.2 μM. Compounds 2, 6 and
with C18 column to yield methyl rosmarinate (6) (3.2 mg), 7 showed moderate activity while 3, 4, 5 and 8 were
oresbiusin B (7) (8.1 mg) and dimethyl lithospermate (8) inactive. Interestingly, all compounds did not affect the
(100 mg). Subfraction F3.2 was isolated over silica gel viability of normal cells (monkey kidney VERO cell
column eluted with dichloromethane-methanol(2:1, v/v) to line) up to 30 μM. Methyl caffeate (2) has been reported
provide rosmarinic acid (5) (3.2 mg). Astragalin (4) (11.0 as a promising anti-cancer agent by inducing apoptosis
mg) was isolated from subfraction F3.3 using C-18 (Balachandran et al., 2015; Kim et al., 2019). In this
column chromatography eluted with methanol-water study, compound 1 showed stronger cytotoxic effect
(1/1, v/v). than methyl caffeate. Further studies are needed to
Compound 1 was isolated as a white amorphous clarify the anti-cancer ability of Ehretia asperula as well
powder and had the molecular formula C15H14O4, as as compound 1.

This work is licensed under a Creative Commons Attribution 4.0 International License. You are free to copy, distribute and perform the work.
You must attribute the work in the manner specified by the author or licensor
 Bangladesh J Pharmacol 2019; 14: 196-197 197

Table I

Cytotoxicity (μM) of the isolated compounds

Compounds Hep-G2 LU-1 HeLa MCF-7 RD Vero
1 7.1 ± 0.7 8.5 ± 1.1 10.2 ± 0.9 >30 7.9 ± 0.8 >30
2 14.5 ± 2.0 >30 17.4 ± 2.16 22.6 ± 2.69 >30 >30
6 22.1 ± 3.6 >30 24.9 ± 2.63 15.1 ± 1.23 >30 >30
7 25.4 ± 3.5 >30 >30 >30 >30 >30
Ellipticine 3.2 ± 0.2 3.0 ± 0.4 1.3 ± 0.1 4.00 ± 0.4 3.3 ± 0.3 >30

This work was supported by Department of Science and A, Badshah A, Rana UA, Noureen Z, Green IR, Draeger S,
Technology, Hoabinh Province, and Vietnam Academy of Schulz B, Krohn K. Coniothyren: A new phenoxyphenyl
Science and Technology (NCVCC30.01/19-19). ether from the endophytic fungus, Coniothyrium sp., J Asian
Nat Prod Res. 2014; 16: 1094-1098.

Dang Dinh Kim1, Vu Thi Nguyet1, Ha Xuan Anh1, Kim KO, Lee D, Hiep NT, Song JH, Lee HJ, Lee D, Kang KS.
Nguyen Thi Thu Trang1, Nguyen Hong Chuyen1, protective effect of phenolic compounds isolated from
Le Mai Huong2, Tran Thi Hong Ha2, Do Hai Ho3, mugwort (Artemisia argyi) against contrast-induced apop-
Nguyen Tien Dat4* tosis in kidney epithelium cell line LLC-PK1. Molecules
1
2019; 24: E195.
Institute of Environmental Technology, Vietnam Academy of Science
and Technology (VAST), 18 Hoang Quoc Viet, Cau Giay, Hanoi, Li L, Peng Y, Yao X, Xu LJ, Wulan T, Liu Y, Shi RB, Xiao PG.
Vietnam; 2Institute of Natural products Chemistry, VAST, 18 Hoang
Quoc Viet, Cau Giay, Hanoi, Vietnam; 3Department of Science and Chemical constituents and biological activities of plants
Technology, Hoabinh, Vietnam; 4Center for research and technology from the genus Ehretia Linn. Chin Herb Med. 2010; 2: 106-11.
transfer, VAST, 18 Hoang Quoc Viet, Cau Giay, Hanoi, Vietnam.
Nguyen TL, Pham TH, Do VT, Huynh TTH. Evaluating the
*Corresponding author: systematic position of Ehretia asperula Zoll. & Moritzi based
email: [email protected] on ITS1, matK and trnL-trnF DNA sequences. Vietnam J Sci
References Technol Eng. 2017; 59: 61-65.

Balachandran C, Emi N, Arun Y, Yamamoto Y, Ahilan B, Ooike M, Nozawa K, Kawai KI. An epitetrathiodioxo-
Sangeetha B, Duraipandiyan V, Inaguma Y, Okamoto A, piperazine related to emestrin from Emericella foveolata.
Ignacimuthu S, Al-Dhabi NA, Perumal PT. In vitro Photochemistry 1997; 46: 123-26.
anticancer activity of methyl caffeate isolated from Solanum
torvum Swartz. fruit. Chem Biol Interact. 2015; 242: 81-90. Shukla A, Kaur A. A systematic review of traditional uses bio-
active phytoconstituents of genus Ehretia. Asian J Pharm
Hussain H, Kock I, Al-Harrasi A, Abbas G, Rehman NU, Shah Clin Res. 2018; 11: 88-100.