Phytochemistry Letters 15 (2016) 13–15

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Phytochemistry Letters
journal homepage: www.elsevier.com/locate/phytol

Two new anthraquinones with antiviral activities from the barks of

Morinda citrifolia (Noni)
Junfeng Wanga , Xiaochu Qinb , Zhiyun Chena , Zhiran Jua , Weijun Hea,c, Yehui Tana ,
Xiaojiang Zhouc , Zhengchao Tub , Fangguo Luc,* , Yonghong Liua,*
a
CAS Key Laboratory of Tropical Marine Bio-resources and Ecology/Guangdong Key Laboratory of Marine Materia Medica/RNAM Center for Marine
Microbiology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, China
b
Laboratory of Molecular Engineering and Laboratory of Natural Product Synthesis, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of
Sciences, Guangzhou 510530, China
c
School of Medicine, Hunan University of Chinese Medicine, Changsha 410208, China

A R T I C L E I N F O A B S T R A C T

Article history: Two new anthraquinones, 1,3-dihydroxy-5-methoxy-6-methoxymethyl-2-methyl-9,10-anthraquinone

Received 1 September 2015 (1) and 1,3-dihydroxy-5-methoxy-2,6-bismethoxymethyl-9,10-anthraquinone (2), together with ten
Received in revised form 31 October 2015 known anthraquinone derivatives (3–12), three coumarin derivatives (13–15), and 6-gingerol (16) were
Accepted 9 November 2015
isolated from the barks of Morinda citrifolia (Noni) collected in the Yongxing island of Xisha. The
Available online 21 November 2015
structures of compounds (1–16) were determined on the basis of extensive spectroscopic analyses, as
well as by comparison with literature reports. The new compounds 1 and 2 were tested for their antiviral,
Keywords:
cytotoxic, and antibacterial activities. In the primary bioassays, compounds 1 and 2 displayed weak anti-
Morinda citrifolia
Anthraquinone
H1N1 activity with IC50 values of 66.1 and 10.5 mM, respectively. In addition, compound 2 showed weak
Antiviral activity anti-H3N2 activity with IC50 value of 11.5 mM, and had weak antimicrobial activity against Staphylococcus
Antimicrobial activity aureus with MIC value of 24.5 mM.
ã 2015 Phytochemical Society of Europe. Published by Elsevier B.V. All rights reserved.

1. Introduction bismethoxymethyl-9,10-anthraquinone (2) (Fig. 1), together with

Morinda citrifolia L., commonly known as “noni”, belongs to the
Rubiaceae family. This plant is native from South-East Asia to
Australia, and has been used by the Polynesians for more than
2000 years as a food, dye, and herbal remedies to cure several
diseases (Wang et al., 2002). In South-East Asia, the fruits are used
to treat diabetes, as well as for treating swollen spleen, liver
diseases and cough (Ee et al., 2009). The roots and barks of “noni”
are well known for their abundance of anthraquinones (Inoue et al.,
1981; Siddiqui et al., 2006; Ee et al., 2009). Anthraquinones are
known to have a variety of interesting biological profiles, such as
antiviral, antioxidant, antimicrobial, cytotoxic, anti-inflammatory,
and anti-osteoporotic activities (Akihisa et al., 2007; Wu et al.,
2009; Locatelli, 2011; Kremer et al., 2012; Locatelli et al., 2012;
Zheng et al., 2012; Zengin et al., 2015). In our continuing
investigations on novel structures from “noni”, two new anthra-
quinones, 1,3-dihydroxy-5-methoxy-6-methoxymethyl-2-methyl-
9,10-anthraquinone (1) and 1,3-dihydroxy-5-methoxy-2,6-
* Corresponding authors. Fax: +86 020 89023244.
E-mail addresses: lufangguo0731@163.com (F. Lu), yonghongliu@scsio.ac.cn
(Y. Liu).
ten known anthraquinone derivatives, 1,5,15-trimethylmorindol
(3) (Takashima et al., 2007), morindone-6-methyl-ether (4) (Ee
et al., 2009), 1-hydroxy-5-methoxyanthraquinone (5) (Chang and
Lee, 1985), 1,4-dimethoxyl-2-hydroxyanthraquinone (6) (Li et al.,
2012), 2,3-dihydroxyl-1-methoxyl-anthraquinone (7) (Li et al.,
2012), rubiadin (8) (Yoo et al., 2010), lucidin 3-methyl ether (9)
(Fraga et al., 2009), lucidin 1,3-dimethyl ether (10) (Fraga et al.,
2009), damnacanthol (11) (Li et al., 2006), digiferruginol (12)
(Zhang et al., 2010), scopoletin (13) (Wu et al., 2009), fraxidin (14)
(Yasuda et al., 2006), isofraxidin (15) (Tian et al., 2008), 6-gingerol
(16) (Lee et al., 2001) were isolated. Herein, we report the isolation,
structure elucidation and bioactivities of the new compounds (1
and 2) from the barks of M. citrifolia.
2. Results and discussion
Compound (1) was obtained as yellow powder with the
molecular formula C18H16O6 as determined by HRESIMS at m/z
329.1016 [M + H]+ (calcd 329.1020), indicating 11 degrees of
unsaturation. The 1H NMR spectrum contained signals attributable
to three aromatic protons, a pair of doublets (J = 7.4 Hz) due to two
ortho-positioned aromatic protons (H-7 and H-8), and a singlet

http://dx.doi.org/10.1016/j.phytol.2015.11.006
1874-3900/ ã 2015 Phytochemical Society of Europe. Published by Elsevier B.V. All rights reserved.
14 J. Wang et al. / Phytochemistry Letters 15 (2016) 13–15

Fig. 1. The chemical structures of the new compounds 1 and 2.

(dH 7.16) due to one isolated aromatic proton (H-4). This spectrum (DEPT) spectrum, another methoxymethyl group at C-2 was
also showed signals for four substituents, which were assignable to observed in 2, instead of the methyl group (2-CH3). This deduction
a methyl, an O-methyl group, a methoxymethyl group, and a was further supported by the HMBC correlation of H2-11 with C-1,
phenolic hydroxyl peri to the carbonyl group. The 13C NMR C-2, and C-3, and 11-OCH3 with C-11 (Fig. 2). Therefore, compound
spectrum of 1 displayed signals for all 18C-atoms, including those 2 was identified as 1,3-dihydroxy-5-methoxy-2,6-bismethoxy-
of the anthraquinone skeleton (dC 185.5, 180.9, 163.5, 162.0, 157.9, methyl-9,10-anthraquinone.
140.4, 134.2, 133.6, 133.1, 124.8, 122.4, 116.4, 108.3, 107.4), one The new compounds 1 and 2 were evaluated for their antiviral,
methyl group (dC 8.0), two O-methyl groups (dC 61.6, 58.2), and one antibacterial, and cytotoxic activities. In the primary bioassays,
methylene group (dC 68.3). The connectivity of the protons and C- compounds 1 and 2 displayed weak anti-H1N1 activity with IC50
atoms was established by the 1H, 13C, HMQC spectrum (Table 1). values of 66.1 and 10.5 mM, respectively. In addition, compound 2
The HMBC correlation of CH3 (dH 2.04) with C-1, C-2, and C- showed weak anti-H3N2 activity with IC50 value of 11.5 mM, and
3 suggested that the CH3 group should be located at C-2. The had weak antimicrobial activity against Staphylococcus aureus with
location of the OCH3 (dH/C 3.81/61.6) at C-5 was supported by MIC value of 24.5 mM. But none of the compounds exhibited
HMBC correlation of OCH3 (dH 3.81) with C-5 (Fig. 2). The location cytotoxic effects on the ten cancer cell lines (K562, U937, MOLT-4,
of the methoxymethyl group at C-6 was supported by HMBC HL60, Hela, DU145, MCF-7, A549, SGC-7901, and H1975).
correlation of CH2OCH3 (dH 4.57) with C-6 (Fig. 2). One OH group
located at C-1 was confirmed by HMBC correlations of the OH H- 3. Experimental
atom signal (dH 13.04) with C-1, C-1a, and C-2, indicating a
phenolic H-atom that is hydrogen-bonded to the neighboring CO 3.1. General experimental procedures
group. Another phenolic OH group, of which the H-atom signal was
undetected, should be located at C-3. Thus, compound 1 was Optical rotations were measured with a PerkineElmer 341 po-
identified as 1,3-dihydroxy-5-methoxy-6-methoxymethyl-2- larimeter. UV spectra were recorded on a Shimadzu UV-2401PC
methyl-9,10-anthraquinone. spectrometer. 1H, 13C NMR, DEPT, and 2D-NMR spectra were
Compound (2) was deduced to have the molecular formula recorded on the Bruker DRX-500 spectrometer using TMS as
C19H18O7 with 11 degrees of unsaturation, having one CH2O unit internal standard and chemical shifts were recorded as d-values.
more than 1, on the basis of HRESIMS data (m/z 359.1114 [M + H]+). HRESIMS (including ESIMS) spectra were recorded on an Applied
Detailed analysis of the 1D- and 2D-NMR spectra data revealed Biosystems Mariner 5140 spectrometer. TLC and column chroma-
that 2 was very similar to those of 1 (Table 1), indicating that they tography (CC) were performed on plates precoated with silica gel
shared the same skeleton. However, the methyl group (2-CH3) in GF254 (10–40 mm) and over silica gel (200–300 mesh) (Qingdao
the 1H NMR spectrum of 1 disappeared in that of 2. In the 13C-NMR Marine Chemical Factory, China), and Sephadex LH-20 (Amersham
Biosciences, Sweden), respectively. All solvents used were of
analytical grade (Tianjin Fuyu Chemical and Industry Factory).
Table 1 Semipreparative HPLC was performed using an ODS column (YMC-
13
C and 1H NMR spectral data of 1 and 2 (125 and 500 MHz, in DMSO-d6,d in ppm). pack ODS-A, 10  250 mm, 5 mm, 4 mL/min).
Position 1 2
3.2. Plant material
dC Mult dH (J in Hz) dC Mult dH (J in Hz)
1 162.0, s 169.5, s The barks of M. citrifolia (Noni) were collected from the Xisha
1a 108.3, s 106.2, s
Islands of China in November 2013. The species was identified by
2 116.4, s 115.6, s
3 163.5, s 164.1, s Dr. Zhiyun Chen (one of our authors). And a voucher specimen (No.
4 107.4, d 7.16, s 110.7, d 7.03, s S201301) was deposited at the CAS Key Laboratory of Tropical
4a 133.1, s 133.6, s Marine Bio-resources and Ecology, South China Sea Institute of
5 157.9, s 157.9, s
Oceanology, Chinese Academy of Sciences.
6 140.4, s 139.6, s
7 133.6, d 7.82, d, (7.4) 133.6, d 7.81, d, (7.8)
8 122.4, d 8.00, d, (7.4) 122.2, d 8.00, d, (7.8) 3.3. Extraction and isolation
9 185.5, s 183.3, s
9a 134.2, s 135.0, s The dried and powdered barks of M. citrifolia (Noni) (8.5 kg)
10 180.9, s 181.7, s
were extracted with 80% EtOH under reflux for 3  3 h. Removal of
10a 124.8, s 124.8, s
11 8.0, q 2.04, s 61.4, t 4.40, s the solvent in vacuo afforded a brown residue, which was
12 68.0, t 4.57, s 68.3, t 4.56, s suspended in H2O followed by successive partition with EtOAc
5-OCH3 61.6, q 3.81, s 61.6, q 3.79, m and n-BuOH. The EtOAc extract (63 g) was subjected to column
11-OCH3 57.4, q 3.17, s
chromatography (CC) over silica gel (200–300 mesh) eluting with
12-OCH3 58.2, q 3.40, s 58.2, q 3.39, s
1-OH 13.04, s 13.44, s
gradient CHCl3/MeOH (1:0–0:1) to afford seven fractions based on
TLC properties. Fraction 4 was passed through Sephadex LH-20
 J. Wang et al. / Phytochemistry Letters 15 (2016) 13–15
Fig. 2. The key HMBC correlations of 1 and 2.
(MeOH) to obtain six fractions (Frs. 4-1-4-6). Fr. 4-1 was purified by
HPLC (38% CH3CN/H2O) to yield 9 (5.1 mg, tR 8.9 min), 10 (6.4 mg, tR
14.5 min), 3 (5.3 mg, tR 17.1 min), and 16 (14.2 mg, tR 27.7 min),
respectively. Fr. 4-5 was purified by HPLC (72% MeOH/H2O) to yield
2 (18.9 mg, tR 16.9 min). Fraction 5 was directly purified by HPLC
(75% MeOH/H2O) to yield three fractions (Frs. 5-1-5-5), 8 (12.6 mg,
tR 16.8 min), 1 (7.1 mg, tR 18.9 min), and 4 (7.0 mg, tR 20.1 min),
respectively. Fr. 5-1 was purified by HPLC (60% MeOH/H2O) to yield
Fr. 5-1-1, Fr.5-1-2, 11 (6.8 mg, tR 8.9 min), 5 (5.1 mg, tR 12.4 min),
and 12 (9.5 mg, tR 17.2 min), respectively. Fr. 5-1-1 was purified by
HPLC (30% CH3CN/H2O) to yield 14 (7.4 mg, tR 21.8 min), 13
(19.4 mg, tR 23.2 min), and 15 (4.3 mg, tR 26.0 min), respectively.
Similarly, Fr. 5-1-2 was further purified by HPLC (47% CH3CN/H2O)
to yield 7 (2.4 mg, tR 19.2 min) and 6 (4.8 mg, tR 21.6 min),
respectively.
1,3-Dihydroxy-5-methoxy-6-methoxymethyl-2-methyl-9,10-
anthraquinone (1): yellow powder; UV (MeOH) lmax (log e): 206
(4.05), 248 (3.78), 280 (3.86), 410 (3.30) nm; IR (KBr) nmax 3406,
1659, 1628, 1562, 1450, 1431, 1369, 1335, 1285, 1250, 1200, 1115,
1069, 1011 cm 1; 1H NMR and 13C NMR data, see Table 1; HRESIMS
m/z 329.1016 [M + H]+ (calcd for C18H17O6, 329.1020).
1,3-Dihydroxy-5-methoxy-2,6-bismethoxymethyl-9,10-an-
thraquinone (2): yellow powder; UV (MeOH) lmax (log e): 208
(4.11), 248 (3.98), 281 (3.94), 417 (3.44) nm; IR (KBr) nmax 3395,
1667, 1624, 1589, 1450, 1369, 1331, 1288, 1254, 1196, 1099, 1065,
1018 cm 1; 1H NMR and 13C NMR data, see Table 1; HRESIMS m/z
359.1114 [M + H]+ (calcd for C19H19O7, 359.1125).
Acknowledgments
This work was supported financially by the National Key Basic
Research Program of China (973)s Project (2011CB915503), the
National Natural Science Foundation of China (Nos. 21502204,
31270402, 21172230, and 41476135), the Strategic Priority
Research Program of the Chinese Academy of Sciences
(XDA11030403), Guangdong Marine Economic Development and
Innovation of Regional Demonstration Project (GD2012-D01-001),
the Aid program for Science and Technology Innovative Research
Team in Higher Educational Instituions of Hunan Province (No. 15,
Grxjb-4).
Appendix A. Supplementary data
Supplementary data associated with this article can be found,
in the online version, at http://dx.doi.org/10.1016/j.phytol.
2015.11.006.
15
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