Dalvelutinoside, a new isoflavone glycoside from the methanol extract of Dalbergia velutina roots

Article abstract of DOI:10.1080/14786419.2015.1114936

A new isoflavone glycoside, dalvelutinoside (1), together with one known isoflavone (2) and five known isoflavone glycosides (3–7) were isolated from the methanol extract of the roots of Dalbergia velutina. Their structures were determined by spectroscopic analysis. All isolated compounds were evaluated for their cytotoxicity against KB and HeLa cell lines.

Full text of DOI:10.1080/14786419.2015.1114936

Natural Product Research
Formerly Natural Product Letters
ISSN: 1478-6419 (Print) 1478-6427 (Online) Journal homepage: http://www.tandfonline.com/loi/gnpl20
Dalvelutinoside, a new isoflavone glycoside from
the methanol extract of Dalbergia velutina roots
Sutin Kaennakam, Pongpun Siripong & Santi Tip-pyang
To cite this article: Sutin Kaennakam, Pongpun Siripong & Santi Tip-pyang (2015):
Dalvelutinoside, a new isoflavone glycoside from the methanol extract of Dalbergia velutina
roots, Natural Product Research, DOI: 10.1080/14786419.2015.1114936
To link to this article: http://dx.doi.org/10.1080/14786419.2015.1114936
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Date: 26 November 2015, At: 03:45

Natural Product research, 2015
ꢁꢁp://ꢂx.ꢂꢃi.ꢃꢄg/10.1080/14786419.2015.1114936
ꢀ
Dalvelutinoside, a new isoflavone glycoside from the
methanol extract of Dalbergia velutina roots
a
b
a
Sutin Kaennakam , Pongpun Siripong and Santi Tip-pyang
ꢅ
Fꢅꢆꢇꢈꢁy ꢃf sꢆiꢉnꢆꢉ, dꢉpꢅꢄꢁmꢉnꢁ ꢃf cꢀꢉmiꢊꢁꢄy, Nꢅꢁꢇꢄꢅꢈ Pꢄꢃꢂꢇꢆꢁꢊ rꢉꢊꢉꢅꢄꢆꢀ uniꢁ, cꢀꢇꢈꢅꢈꢃngkꢃꢄn univꢉꢄꢊiꢁy,
b
Bꢅngkꢃk, tꢀꢅiꢈꢅnꢂ; rꢉꢊꢉꢅꢄꢆꢀ diviꢊiꢃn, Nꢅꢁꢇꢄꢅꢈ Pꢄꢃꢂꢇꢆꢁꢊ rꢉꢊꢉꢅꢄꢆꢀ sꢉꢆꢁiꢃn, Nꢅꢁiꢃnꢅꢈ cꢅnꢆꢉꢄ Inꢊꢁiꢁꢇꢁꢉ, Bꢅngkꢃk,
tꢀꢅiꢈꢅnꢂ
ABSTRACT
ARTICLE HISTORY
rꢉꢆꢉivꢉꢂ 30 Jꢇꢈy 2015
aꢆꢆꢉpꢁꢉꢂ 12 sꢉpꢁꢉmbꢉꢄ
A new isoflavone glycoside, dalvelutinoside (1), together with one
known isoflavone (2) and five known isoflavone glycosides (3–7) were
isolated from the methanol extract of the roots of Dalbergia velutina.
Their structures were determined by spectroscopic analysis. All
isolated compounds were evaluated for their cytotoxicity against KB
and HeLa cell lines.
2015
KEYWORDS
dꢅꢈvꢉꢈꢇꢁinꢃꢊiꢂꢉ; Dalbergia
velutina; iꢊꢃflꢅvꢃnꢉ
gꢈyꢆꢃꢊiꢂꢉ; ꢆyꢁꢃꢁꢃxiꢆiꢁy
1
. Introduction
Dalbergia velutina is a perennial climbing tree (Leguminosae), mainly distributed in the
north-eastern ofThailand. The previous phytochemistry of this genus resulted in the isolation
of various compounds classified as isoflavones (Nagarajan et al. 2006), isoflavanes, isoflava-
nones, neoflavones, pterocarpans, anthraquinones, triterpenes, cinnamyl esters (Vasudeva
et al. 2009) and phenolic compounds (Inui et al. 2014). Many of these compounds have
demonstrated several interesting biological activities such as anticancer, antimicrobial, anti-
oxidant, antiulcerogenic, antiplasmodial (Zheng et al. 2012 and Mutai et al. 2013), analgesic,
anti-inflammatory and antibacterial (Narayanan et al. 2007).
In our previous studies on D. velutina (Kaennakam et al. 2015), we reported six pterocar-
pans with cytotoxicity against KB and HeLa cell lines from CH Cl extract of the roots of this
2
2
plant. Our continuous study on the methanol extract of this plant led to the isolation of a
new isoflavone glycoside, dalvelutinoside (1) along with one known isoflavone, olibergin
CONTACT sꢅnꢁi tip-pyꢅng
ꢊꢅnꢁi.ꢁi@ꢆꢀꢇꢈꢅ.ꢅꢆ.ꢁꢀ
sꢇppꢈꢉmꢉnꢁꢅꢈ ꢂꢅꢁꢅ fꢃꢄ ꢁꢀiꢊ ꢅꢄꢁiꢆꢈꢉ ꢆꢅn bꢉ ꢅꢆꢆꢉꢊꢊꢉꢂ ꢅꢁ ꢀꢁꢁp://ꢂx.ꢂꢃi.ꢃꢄg/10.1080/14786419.2015.1114936.
©
2015 tꢅyꢈꢃꢄ & Fꢄꢅnꢆiꢊ

2
S. KAꢀNNAKAM ꢀT AL.
R1
R O
7
9 O
C
2
HO
O
O
1
OCH3
A
1
'
3'
R2
5
10
4
B
OH
O
OH
6'
R3
OH
OCH3
R2
R1
-Glc
H
-Apio(1
-Apio(1
R2
R3
R1
-Glc
-Glc OH
R2
1
2
3
4
H
H
H
OH
OH
OH
5
6
7
H
6)- -Glc
-Glc OCH3
6)- -Glc OCH3 OCH3
OH
OH
O
O
O
O
OH
-Apio(1
6)- -Glc =
OH
HO
OH
Figure 1. tꢀꢉ ꢊꢁꢄꢇꢆꢁꢇꢄꢉ ꢃf ꢆꢃmpꢃꢇnꢂꢊ 1–7.
Table 1. In vitro ꢆyꢁꢃꢁꢃxiꢆiꢁy ꢃf ꢆꢃmpꢃꢇnꢂꢊ 1–7 ꢅgꢅinꢊꢁ KB ꢅnꢂ hꢉlꢅ ꢆꢉꢈꢈ ꢈinꢉꢊ.
IC50 (μM)
IC50 (μM)
Compound
KB
HeLa
Compound
KB
HeLa
1
>100
48.06
>100
0.22
>100
59.82
>100
0.10
4
5
6
7
63.77
>100
>100
>100
86.36
>100
>100
>100
2
3
aꢂꢄiꢅmyꢆin
A (2) (Ito et al. 2003) and five known isoflavone glycosides, 2′,5′-dimethoxy-genistein
7
7
-O-β-D-apiofuranosyl-(1‴→6″)-O-β-D-glucopyranoside (3) (Marzouk et al. 2008), caviunin
-O-β-D-apiofuranosyl-(1‴→6″)-β-D-glucopyranoside (4) (Dixit et al. 2012), genistein-8-C-β-
D-glucopyranoside(5), orobol-8-C-β-D-glucopyranoside (6) (Zapesochnaya & Laman 1978)
and dalpanitin (7) (Adinarayana & Rajasekhara 1972) (Figure 1). In addition, cytotoxicity result
against KB and HeLa cells of all isolated compounds are also reported. Their structures were
identified by interpretation of their spectroscopic data as well as comparison with those
reported in the literature.
2
. Results and discussion
The MeOH extract from the roots of D. velutina was separated by silica gel (Merck Art 7730)
and Sephadex LH-20 column chromatography to obtain the compounds 1–7. Their structures
were elucidated on the basis of detailed spectroscopic analysis including 1D and 2D NMR
spectroscopy and mass spectrometry techniques.
2
0
Dalvelutinoside (1) was obtained as a white amorphous powder,[ꢀ]D −62.8 (c 1.0, DMSO).
Its molecular formula was determined as C H O by HRꢀSIMS measurement through the
2
3
24 12
+
pseudomolecular ion peak at m/z 515.1161 [M + Na] (calcd. for C H O Na, 515.1165).
2
3
24 12
The UV spectrum displayed absorption bands at λmax 244, 262 and 309 nm. The IR spectrum

NATURAL PRODUCT RꢀSꢀARCH
3
−
1
−1
showed absorption bands for hydroxyl groups (3410 cm ), a carbonyl group (1665 cm )
−1
1
and aromatic moieties (1540 and 1500 cm ). The H NMR spectrum had signals of four
aromatic protons at δ 6.47 (1H, d, J = 2.0 Hz, H-6), 6.59 (1H, s, H-3′), 6.72 (1H, d, J = 2.4 Hz,
H
H-8), 6.88 (1H, s, H-6′), two phenolic protons at δ 9.25 (1H, s, OH-4′) and 12.92 (1H, s, OH-5,
H
chelating to the carbonyl) which were consistent with a 5,7-disubstituted aromatic ring A and
a 2,4,5-trisubstituted aromatic ring B. The four hydroxyl groups of sugar unit at δ 4.61 (1H,
H
t, J = 5.4 Hz, OH-6″), 5.06 (1H, d, J = 5.6 Hz, OH-3″), 5.13 (1H, d, J = 4.4 Hz, OH-4″), 5.41 (1H,d,
J = 4.8 Hz, OH-2″), and two methoxy at δ 3.64 (3H, s, OCH ′) and 3.72 (3H, s, OCH ′). In
H
3–2
3–5
addition, one olefinic proton at δ 8.28 (1H, s, H-2) is the characteristic of isoflavone skeleton.
H
1
3
The C NMR spectra showed 23 signals comprising 2 methyl, 1 methylene, 10 methine and
1
0 quaternary carbons. Compound 1 was isoflavone glucoside with anomeric proton signal
at δ 5.06 (1H, d, J = 7.2 Hz, H-1″) and anomeric carbon resonance at δ 99.7 (C-1″) of glucose
H
C
indicated the β-configuration. In the HMBC correlations of 1 (Figure S6), the anomeric proton
at H-1″ showed cross peaks with C-3″ (δ 77.0) and C-7 (δ 162.7), hydroxy proton at δ 12.92
C
C
H
(
1H, s, OH-5) had the correlations with C-5 (δ 161.3), C-6 (δ 99.3) and C-10 (δ 105.7) and
C C C
aromatic proton at δ 6.72 (1H, d, J = 2.4 Hz, H-8) correlated with C-7 (δ 162.7), C-9 (δ 157.0)
H
C
C
and C-10. The above data indicated that one glucoside unit was located at C-7 of ring A, while
those H-2 to C-3 (δ 120.4), C-4 (δ 180.1), C-9 (δ 157.0) and C-1′ (δ 108.8) established that
C
C
C
C
ring C was attached to C-1′ of ring B and two methoxyl groups connected to C-2′ and C-5′
showed cross peak with δ 151.8 and 140.9, respectively. In addition, a hydroxyl group at
C
1
13
position 4′ related to C-3′ (δ 100.7), C-4′ (δ 147.7) and C-5′ (δ 140.9). The H and C NMR
spectroscopic data were shown to be quite similar to those of the known isoflavone gly-
C
C
C
coside, 2′,5′-dimethoxy-genistein 7-O-β-D-apiofuranosyl-(1‴→6″)-O-β-D-glucopyranoside
(
3), except for compound 1 comprised only one glucose unit. The appearance of a glu-
copyranosyl group was confirmed by TLC analysis of the hydrolysis of 1 with 1 M HCl, and
the absolute configuration of this sugar moiety was identified as D-glucopyranoside by
HPLC analysis. Thus, the complete assignment of dalvelutinoside (1) was determinated as
2
′,5′-dimethoxy-genistein 7-O-β-D-glucopyranoside. The in vitro cytotoxic activities of these
compounds are shown in Table 1, compounds 2 and 4 showed weak cytotoxicity against
KB and HeLa cells with IC values of 48.06, 63.77 μM and 59.82, 86.36 μM, respectively, and
5
0
other compounds were inactive.
3
. Experimental
.1. General experimental procedures
D and 2D NMR spectra were recorded on a Bruker 400 AVANCꢀ spectrometer. Melting points
3
1
were obtained using Fisher-Johns Melting Point apparatus. HRꢀSIMS spectra were obtained
using a Bruker MICROTOF model mass spectrometer. IR data were obtained using a Nicolet
6700 FT-IR spectrometer using KBr discs. UV-visible absorption spectra were taken on a
UV-2550 UV–vis spectrometer (Shimadzu, Kyoto, Japan). Optical rotation was recorded by
Jasco P-1010 Polarimeter. HPLC analysis were obtained using a Alltech System equipped with
model 626 binary gradient pumps, selectTM degasser, 580 autosampler, 2000 ꢀS evaporative
light scattering detector (Alltech) and peak simple chromatography data system software.

4
S. KAꢀNNAKAM ꢀT AL.
3
.2. Plant material
The roots of D. velutina were collected from Sahatsakhan district, Kalasin province, Thailand,
in October 2014. The plant material was identified by Ms Suttira Khumkratok, and a voucher
specimen was deposited as a reference (Khumkratok No. 4-12) at the Walai Rukhavej Botanical
Research Institute, Mahasarakham University.
3
.3. Extraction and isolation
The air-dried roots of D. velutina (5.5 kg) were extracted with CH Cl and MeOH over a period
2
2
of 3 days at room temperature, respectively (3 × 15 L). Removal of the solvent under reduced
pressure provided MeOH (80.5 g) crude extracts. The MeOH crude extract was further sepa-
rated by column chromatography over silica gel (Merck Art 7730) and eluted with a gradient
of ꢀtOAc-MeOH (100% ꢀtOAc, 90, 80 and 70% ꢀtOAc-MeOH each 5 L, respectively) to give
five fractions (A–ꢀ). Fraction A (4.5 g) was purified by Sephadex LH-20 column (150 g) with
100% MeOH (1.5 L) to afford compound 1 (12.5 mg) and compound 2 (20.5 mg). Fraction
B (6.5 g) was separated by Sephadex LH-20 column (150 g) eluted with 100% MeOH (1.5 L)
to give compound 5 (8.5 mg), compound 6 (10.5 mg) and compound 7 (14.5 mg). Finally,
fraction C (5.0 g) was subjected to a Sephadex LH-20 column (150 g) eluted with 100% MeOH
(
1 L) to yield compound 3 (15.0 mg) and compound 4 (16.5 mg).
Dalvelutinoside (1): White amorphous powder; m.p. 218.0–219.0 °C; [ꢀ] −62.8 (c 1.0,
2
0
D
−1 1
DMSO); UV λmax (DMSO) 244, 262, 309 nm; IR νmax (KBr): 3410, 1665, 1540, 1500 cm ; H NMR
(
400 MHz, in DMSO-d ) δ : 12.92 (1H, s, OH-5), 9.25 (1H, s, OH-4′), 8.28 (1H, s, H-2), 6.88 (1H,
6
H
s, H-6′), 6.72 (1H, d, J = 2.4 Hz, H-8), 6.59 (1H, s, H-3′), 6.47 (1H, d, J = 2.0 Hz, H-6), 5.41 (1H, d,
J = 4.8 Hz, OH-2″), 5.13 (1H, d, J = 4.4 Hz, OH-4″), 5.07 (1H, d, J = 5.6 Hz, OH-3″), 5.06 (1H, d,
J = 7.2 Hz, H-1″), 4.61 (1H, t, J = 5.4 Hz,OH-6″), 3.72 (3H, s, OCH ′), 3.70 (1H, m, H-6″), 3.64
3
–5
(
3H, s, OCH ′), 3.46 (1H, m, H-6″), 3.45 (1H, m, H-3″), 3.32 (1H, m, H-5″), 3.29 (1H, m, H-2″),
3
–2
13
3
1
1
6
.18 (1H, m, H-4″). C NMR (100 MHz, in DMSO-d ) δ : 180.1 (C-4), 162.7 (C-7), 161.3 (C-5),
6
C
57.0 (C-9), 155.7 (C-2), 152 (C-2′), 147.7 (C-4′), 141 (C-5′), 120.4 (C-3), 116.4 (C-6′), 108.8 (C-1′),
05.7 (C-10), 100.7 (C-3′), 99.7 (C-1″), 99.3 (C-6), 94.4 (C-8), 77.0 (C-3″), 76.2 (C-5″), 72.9 (C-2″),
+
9.4 (C-4″), 60.4 (C-6″), 56.5 (C-5′, OCH ), 55.8 (C-2′, OCH ); HRꢀSIMS m/z: 515.1161 [M + Na]
3
3
(
calcd. for C H O Na, 515.1165).
23 24 12
3
.4. Acidic hydrolysis and HPLC analysis
A solution of dalvelutinoside (1) (2 mg) in 1 M HCl (1.0 mL) was heated at reflux for 1 h and
then the reaction mixture was neutralised with an equal volume of 1 M NaOH and extracted
with CH Cl (5 mL). The sugar moiety was identified as glucose by co-TLC analysis (ꢀtOAC:
2
2
MeOH: H O, 1:8:1) of the aqueous solution in comparison with an authentic glucose. In
2
addition, the glucose was identified as D-glucose by HPLC analysis (column: lichrocart-NH2
(
250 × 4.0 mm), carrier: 82% ACN in H O (1.5 mL/min), retention time: 8.133 min) in compar-
2
ison with an authentic D-glucose.
3
.5. Cytotoxicity assay
All isolated compounds (1–7) were subjected to cytotoxic evaluation against KB (human
epidermoid carcinoma) and HeLa (human cervical carcinoma) cell lines employing the

NATURAL PRODUCT RꢀSꢀARCH
5
colorimetric method (Skehan et al. 1990; Kongkathip et al. 2003; Kaennakam et al. 2015).
Adriamycin was used as the reference substance which exhibited activity against KB and
HeLa cell lines.
4
. Conclusion
The MeOH crude extract from the roots of D. velutina comprises one new dalvelutinoside
1), one known isoflavone (2) and five known isoflavone glycosides (3–7). Compound 3 was
(
isolated for the first time from this genus. Compounds 2 and 4 showed weak cytotoxicity
against KB and HeLa cells with IC values of 48.06, 63.77 μM and 59.82, 86.36 μM, respec-
5
0
tively. Therefore, we believe that this plant is an important source for the diverse structure
of isoflavone glycosides and should be further investigated for other biological activities.
Supplementary material
Supplementary material relating to this paper is available online, along with Figures S1–S6.
Acknowledgements
We also thank Ms. Suttira Khumkratok, Walai Rukhavej Botanical Research Institute, Mahasarakham
University, Mahasarakham 44000, Thailand, for identification and deposition of the plant material.
Disclosure statement
No potential conflict of interest was reported by the authors.
Funding
This work was supported by the 90th anniversary of Chulalongkorn University Fund (Rat chadaphisek-
somphot ꢀndowment Fund).
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