A new flavonoid glucoside from Rhododendron seniavinii

Article abstract of DOI:10.1080/10286020.2014.989222

The leaves of Rhododendron seniavinii Maxim with little phytochemical information are used as folk remedies for the treatment of acute and chronic bronchitis in China. In our pursuing for the biologically active chemical constituents in the leaves, a new flavonoid glycoside 5,7,3′-trimethoxy-quercetin-3-O-β-D-glucopyranoside (1) was isolated from the water extract of its leaves, together with two known compounds 5,7,3′-trimethoxy-quercetin (2) and ovafolinin B-9′-O-β-D-glucopyranoside (3). The structures of the new flavonoid glucoside as well as two known compounds were elucidated by spectroscopic and chemical methods.

Full text of DOI:10.1080/10286020.2014.989222

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A new flavonoid glucoside from
Rhododendron seniavinii
a
a
a
a
Qing-Qing Wang , Chun Wu , Ying Zhang , Bai-Lian Liu & Guang-
a
Xiong Zhou
a
Guangdong Province Key Laboratory of Pharmacodynamic
Constituents of Traditional Chinese Medicine and New Drugs
Research, Institute of Traditional Chinese Medicine and
Natural Products, College of Pharmacy, Jinan University,
Guangzhou510632, China
Published online: 10 Dec 2014.
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To cite this article: Qing-Qing Wang, Chun Wu, Ying Zhang, Bai-Lian Liu & Guang-Xiong Zhou
2014): A new flavonoid glucoside from Rhododendron seniavinii, Journal of Asian Natural Products
(
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Journal of Asian Natural Products Research, 2014
http://dx.doi.org/10.1080/10286020.2014.989222
A new flavonoid glucoside from Rhododendron seniavinii
Qing-Qing Wang, Chun Wu, Ying Zhang, Bai-Lian Liu and Guang-Xiong Zhou*
Guangdong Province Key Laboratory of Pharmacodynamic Constituents of Traditional Chinese
Medicine and New Drugs Research, Institute of Traditional Chinese Medicine and Natural Products,
College of Pharmacy, Jinan University, Guangzhou 510632, China
(
Received 18 July 2014; final version received 14 November 2014)
The leaves of Rhododendron seniavinii Maxim with little phytochemical information
are used as folk remedies for the treatment of acute and chronic bronchitis in China.
In our pursuing for the biologically active chemical constituents in the leaves, a new
0
flavonoid glycoside 5,7,3 -trimethoxy-quercetin-3-O-b-D-glucopyranoside (1) was
isolated from the water extract of its leaves, together with two known compounds
0
0
5
,7,3 -trimethoxy-quercetin (2) and ovafolinin B-9 -O-b-D-glucopyranoside (3). The
structures of the new flavonoid glucoside as well as two known compounds were
elucidated by spectroscopic and chemical methods.
0
Keywords: Rhododendron seniavinii; flavonoid glucoside; 5,7,3 -trimethoxy-querce-
tin-3-O-b-D-glucopyranoside
1
.
Introduction
compounds [3]. Herein, we report the
isolation and structural elucidation of the
new compound.
Rhododendron seniavinii Maxim (Erica-
ceae), a traditional medicinal plant, is
widely distributed in the southern pro-
vinces of China. This plant is locally
known as “Man Shang Bai,” and its leaves
have been used as folk medicine for the
treatment of acute and chronic bronchitis
2
.
Results and discussion
Compound 1 was obtained as a light
yellow amorphous powder. Its molecular
formula was determined to be C H O
4 26 12
[1]. Several Chinese patent drugs contain-
2
ing this plant, such as Man-Shang-Bai
syrup, were produced and sold now.
Although pharmacological studies on this
plant had demonstrated its effects on
respiratory system and analgesic and
anti-inflammatory aspects, there were few
reports about its chemical constituents [2].
In order to better understand the biological
substances existed in the leaves, the
chemical constituents of the leaves from
the titled plant were investigated, and one
new flavonoids glycoside (Figure 1)
together with two known compounds
were isolated from the water-soluble
extract of the leaves of R. seniavinii,
except for a series of other known
by HR-ESI-MS, which showed
þ
a
[M þ H] peak at m/z 507.1496. The IR
spectra showed a strong absorption band at
2
1
1631 cm
group, and an intense broad band centered
for a conjugated carbonyl
2
1
at 3383 cm from the vibration (nOH) of
hydroxyl groups. The UV absorption
maxima occurred at 250 nm, implying the
presence of long-conjugated double bond
1
systems in the molecule. The H NMR
spectrum showed signals for three meth-
oxyl groups (d 3.88, 3.84, and 3.84) and
H
five aromatic protons. Among these aro-
matic protons, a set of three protons was
resolved as one ABX spin-coupling
system at dH 7.98 (d, J ¼ 1.8 Hz) for H-
*Corresponding author. Email: guangxzh@sina.com
q 2014 Taylor & Francis

2
Q.-Q. Wang et al.
Figure 1. Chemical structures of compounds 1–3.
0
0
2
, 6.90 (d, J ¼ 8.4 Hz) for H-5 , and 7.53
structure of which was confirmed by
HPLC analysis referenced to the method
[5] reported previously. Thus, the structure
0
(
dd, J ¼ 8.4, 1.8 Hz) for H-6 in a flavonol;
the other AX system at d 6.49 (br s) and
H
0
dH 6.79 (br s) was assigned to H-6 and H-8
protons, respectively. The methoxyl
of 1 was determined as 5,7,3 -trimethoxy-
quercetin-3-O-b-D-glucopyranoside.
0
groups were attached at C-5, 7, and 3 on
the basis of the HMBC correlations
between methyl protons at d 3.88, 3.84,
H
1
13
Table 1. H (300 MHz) and C (75 MHz)
NMR spectral data of compound 1 (d in ppm,
J in Hz).
and 3.84 with carbons at d 160.3 (C-5),
C
0
1
ively. The NOE correlations between H-8
63.9 (C-7), and 146.9 (C-3 ), respect-
1
*
0
and 7-OCH , H-6 and 5-OCH , and H-2
3
3
1
13
0
No.
H
C
and 3 -OCH further confirmed the
3
locations of three methoxyl groups
(Table 1). Hence, the flavonoid part has
the basic structure of a known compound
2
3
4
5
6
7
8
9
10
–
–
–
–
153.2
135.6
172.2
160.3
96.0
0
5,7,3 -trimethoxy-quercetin. The anomeric
proton of the sugar appeared at d 5.42 (d,
6.49 d (2.1)
H
–
6.79 d (2.1)
–
163.9
92.9
J ¼ 7.2 Hz) suggested that it be a glyco-
1
side. The C NMR spectrum (Table 1) of
3
158.2
108.2
121.2
113.4
146.9
149.0
115.1
121.6
101.2
74.5
1
1
6
exhibited signals of six carbons at d
00
–
–
C
0
0
0
00
1
0
01.2 (C-1 ), 77.4 (C-2 ), 74.5 (C-3 ),
0
0
00
00
2
7.98 d (1.8)
–
9.9 (C-4 ), 76.6 (C-5 ), and 60.7 (C-6 ),
0
3
which were in good accordance with the
presence of a glucosyl unit [3]. The
coupling constant value (J ¼ 7.2 Hz) of
the anomeric proton confirmed the b-
linkage of the sugar moiety [4]. A long-
range correlation between the anomeric
0
4
–
0
5
0
6.90 d (8.4)
7.53 dd (8.4, 1.8)
5.42 d (7.2)
3.21–3.23 m
3.22–3.25 m
3.07–3.10 m
3.05–3.09 m
6
0
0
0
0
0
0
1
2
3
4
5
6
5
7
3
76.6
69.9
77.4
00
0
0
0
0
0
0
proton at d 5.42 (d, J ¼ 7.2 Hz, H-1 ) and
H
the carbon at dC 135.6 (C-3) of the
flavonoid skeleton in HMBC spectrum
determined the attachment of the sugar at
C-3 of the aglycone (Figure 2). Acid
hydrolysis of 1 afforded D-glucose, the
3.58 (d, 13.1), 3.36–3.39 (m) 60.7
-OCH
-OCH
3
3
3.84 s
3.88 s
3.84 s
56.2
55.7
56.1
0
-OCH
3
6
Note: Solvent: *DMSO-d .

Journal of Asian Natural Products Research
3
(Agilent Technologies) equipped with a
quaternary pump, a multiple wavelength
detector, an AutoSampler, and an
TM
Ultimate
XB-C18 column (5 mm, 4.6
250 mm, Welch, Potamac, MA, USA)
£
were used for analytical HPLC. Semi-
preparative HPLC was carried out on an
Agilent 1200 Series HPLC instrument
(Agilent Technologies) equipped with a
variable wavelength detector and a
TM
reversed-phase column (Ultimate
XB-
C18, 5 mm, 10 £ 250 mm, Welch). Open
column chromatography (CC) was per-
formed by D101 macroporous resin
(
Tianjin Agricultural Pesticide Corpor-
Figure 2. Key HMBC (H ! C) and NOE
H· · ·H) correlations for structural
determination of compound 1.
ation, Tianjin, China), polyamide plates
(80–100 mesh; Taizhou Luqiao Plaschem-
gel, Taizhou, China), precoated silica-gel
plates (GF254, Yantai Chemical Industry
Research Institute, Yantai, China), ODS
(
Compounds 2 and 3 were obtained as
an amorphous yellow or yellowish pow-
der. Their structures were, respectively,
(50 mm, YMC, Tokyo, Japan), and Sepha-
dex LH-20 (Phamacia Biotech, Zurich,
Switzerland). Thin-layer chromatography
was performed by precoated TLC spraying
with 10% H SO followed by heating. All
0
determined to be 5,7,3 -trimethoxy-quer-
0
cetin [5–7] and ovafolinin B-9 -O-b-D-
glucopyranoside [8] on the basis of NMR
and MS data, as compared with the
literature.
2
4
the reagents were purchased from Tianjin
Damao Chemical Company (Tianjin,
China). Standard sugars D-glucose and L-
glucose, and L-cysteine methyl ester in the
HPLC analysis were purchased from
Adamas-Beta Company (Basel, Switzer-
land). O-Tolyl isothiocyanate was pur-
chased from Sigma Company (Santa
Clara, CA, USA).
3
.
Experimental
3
.1 General experimental procedures
Optical rotations were carried out using a
JASCO P-1020 automatic digital polari-
meter (JASCO Corporation, Tokyo,
Japan). UV spectra were recorded on a
JASCO V-550 UV/VIS spectrometer
3.2 Plant material
(JASCO Corporation). IR spectra were
obtained using a JASCO FT/IR-480 plus
spectrometer (JASCO Corporation) with
KBr pellets. HR-ESI-MS were acquired
using Agilent 6210 LC/MSD TOF mass
spectrometer (Agilent Technologies, Santa
Clara, CA, USA). Measurement of 1D and
The water extract of the leaves of
R. seniavinii was prepared and provided
by the Jiangxi Zixi Pharmaceutical Factory
(Fuzhou, China). The original plant
R. seniavinii and the crude leaf material
from the plant were authenticated by the
technician Mr GJ Lee majored in medic-
inal plants at quality control department in
the factory. The material for extraction
was collected in Zixi County, Jiangxi
Province in 2008 summer. A voucher
specimen with accession no. RS 201001
2
D NMR spectra was run with a Bruker
AV-300 spectrometer (Bruker Corpor-
ation, Fallanden, Switzerland) with TMS
as the internal standard, and chemical
shifts were expressed in d values (ppm).
An Agilent 1200 Series HPLC instrument

4
Q.-Q. Wang et al.
has been deposited in the herbarium of the
College of Pharmacy, Jinan University.
3.4 Acid hydrolysis and HPLC analysis
The absolute configuration of the sugar
moiety in the structures was determined by
themethodofTanakaetal. [5]. Compound1
3
.3 Extraction and isolation
(2 mg) was hydrolyzed with 10ml of 2 M
The water extract (324 g) of the leaves of
R. seniavinii was subjected to D101
macroporous absorption resin column
HCl for 6 h at 908C. The mixture was
evaporated to dryness under vacuum and
then it was extracted by CHCl –H O (1:1)
3
2
(
(
80 £ 20 cm) eluting with EtOH–H O
2
twice to afford the aqueous phase, which
was evaporated to dryness under vacuum.
The residue was dissolved in pyridine (2 ml)
containing L-cysteine methyl ester (2mg)
and heated at 608C for 1 h. Then, O-tolyl
isothiocyanate (10 ml) was added to the
mixture, which was heated at 608C for 1 h.
The reaction mixture was directly analyzed
by reversed-phase HPLC. Analytical HPLC
was performed on the column at 308C with
0 ! 30 ! 50 ! 70 ! 95) to afford five
fractions (Fr A–E). Fr C (15.6 g) was
subjected to CC (column chromatography)
on polyamide eluting with MeOH/H O
2
(
1:4 ! 1:0) to yield 10 fractions (Fr C –
1
C ). Fr C (2.3 g) was subjected to CC
1
0
3
(
ODS, MeOH/H O 1:5 ! 1:0) to yield
2
subfractions Fr C -a–C -g. Fr C -e
3
3
3
(300 mg) was fractionated by CC (Sepha-
dex LH-20, MeOH/H O 50:50) to yield Fr
2
isocratic elution of 25% CH CN containing
3
C -e –C -e . Fr C -e (100 mg) was
4
3
1
3
8
3
0.08% formic acid for 40min and sub-
sequent washing of the column with 90%
CH CN at a flow rate of 1 ml/min. Peaks
finally purified by semi-preparative
HPLC (flow rate; 4.0 ml/min; Rt:
3
1
eluant to obtain compound 1 (15.1 mg).
9.3 min) with CH CN/H O (30:70) as an
3 2
were detected by a UV detector at 250 nm.
The peak of the derivative of1 was observed
at tR 17.940 (D-Glc) min. The standard
sugars D-glucose and L-glucose were
subjected to the same method. The peaks
of standard sugar derivatives were recorded
Fr B -d (58 mg) was finally purified by CC
3
(Sephadex LH-20, MeOH/H O 30:70 as
an eluting solvent) to obtain compound 3
2
(
(
9.8 mg). Fr E (1 g) was subjected to CC
Sephadex LH-20, MeOH) to yield sub-
att 17.816(D-Glc) and 16.459(L-Glc) min.
R
fractions Fr E –E . Fr E (120 mg) was
1
10
7
Following the above procedure, the deriva-
tive of 3 also gave a peak at t 17.910 (D-
Glc) min.
finally purified by semi-preparative HPLC
flow rate; 4.0 ml/min; Rt: 23.5 min) with
MeOH–H O (70:30) as an eluting system
R
(
2
to obtain compound 2 (16.3 mg).
Disclosure statement
No potential conflict of interest was reported by
the authors.
0
3.3.1 5,7,3 -Trimethoxy-quercetin-3-O-
b-D-glucopyranoside (1)
2
D
7
Light-yellow amorphous powder; ½aꢀ
Funding
2
6.7 (c ¼ 0.10, MeOH); UV (MeOH)
This work was financially supported by the the
Chinese National S&T Special Project on
Major New Drug Innovation [grant number
2011ZX09307-002-01].
l
max
nmax: 3383, 1631, 1603, 1517.3, 1457.9,
: 206, 250, 265 (sh) nm; IR (KBr)
1
1
431.1, 1357.9, 1287.1, 1214.9, 1107.9,
2
1
1
065.4, 1030.1, 824.4 cm ; for H NMR
1
3
(
(
300 MHz, DMSO-d ) and C NMR
6
References
75 MHz, DMSO-d ) spectral data, see
6
[
1] Delectis Florae Reipublicae Popularis
Sinicae Agendae Academiae Sinicae
Edita, Florae Reipublicae Popularis Sini-
cae (Science Press, Beijing, 1999), Vol. 57.
þ
Table 1; ESI-MS: m/z 529 [M þ Na] ,
–
05 [M–H] ; HR-ESI-MS: m/z 507.1496
5
þ
[M þ H] (calcd for C H O , 507.1497).
2
4 27 12

Journal of Asian Natural Products Research
5
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