C14-Polyacetylene glucosides from Codonopsis pilosula

Article abstract of DOI:10.1080/10286020.2015.1041932

Seven new C₁₄-polyacetylene glucosides codonopilodiynosides A-G (1-7) were isolated from an aqueous extract of the Codonopsis pilosula roots. Their structures were determined by spectroscopic and chemical methods as (-)-(5S,6E,12E)-tetradeca-6,12-dien-8,10-diyn-1,5,14-triol 5-O-β-d-glucopyranoside (1), (-)-(5S,6E,12E)-tetradeca-6,12-dien-8,10-diyn-1,5,14-triol 5-O-β-d-glucopyranosyl-(1″ → 2′)-β-d-glucopyranoside (2), (-)-(5S,6E,12E)-tetradeca-6,12-dien-8,10-diyn-1,5,14-triol 5,14-di-O-β-d-glucopyranoside (3), (-)-(5S,6E)-tetradeca-6-en-8,10-diyn-1,5,14-triol 5-O-β-d-glucopyranoside (4), (-)-(5S,6E,12E)-tetradeca-6,12-dien-8,10-diyn-1,5-diol 5-O-β-d-glucopyranosyl-(1″ → 2′)-β-d-glucopyranoside (5), (-)-(6S,4E,12E)-tetradeca-4,12-dien-8,10-diyn-1,6-diol 6-O-β-d-glucopyranosyl-(1″ → 2′)-β-d-glucopyranoside (6), and (-)-(5S,6E)-tetradeca-6-en-1,5-epoxy-8,10-diyn-14-ol 14-O-β-d-glucopyranosyl-(1″ → 2′)-β-d-glucopyranoside (7), respectively. The absolute configurations of 1-7 were assigned by enzymatic hydrolysis followed by isolation of glucose and aglycones (1a and 4a-7a), and subsequent comparison of specific rotation, TLC, and ¹H NMR data of the glucose with an authentic sugar sample and application of modified Mosher's method based on the MPA determination rule of Δδ^RS values for 1a and 4a, and Δδ^S values for 6a. The configuration of 7 was assigned by electronic circular dichroism calculations based on the quantum-mechanical time-dependent density functional theory.

Full text of DOI:10.1080/10286020.2015.1041932

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C14-Polyacetylene glucosides from
Codonopsis pilosula
Yue-Ping Jianga, Yu-Feng Liua, Qing-Lan Guoa, Zhi-Bo Jianga,
Cheng-Bo Xua, Cheng-Gen Zhua, Yong-Chun Yanga, Sheng Lina &
Jian-Gong Shia
a State Key Laboratory of Bioactive Substance and Function of
Natural Medicines, Institute of Materia Medica, Chinese Academy
of Medical Sciences and Peking Union Medical College, Beijing
100050, China
Published online: 26 May 2015.
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To cite this article: Yue-Ping Jiang, Yu-Feng Liu, Qing-Lan Guo, Zhi-Bo Jiang, Cheng-Bo Xu, Cheng-
Gen Zhu, Yong-Chun Yang, Sheng Lin & Jian-Gong Shi (2015): C14-Polyacetylene glucosides from
Codonopsis pilosula, Journal of Asian Natural Products Research
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Journal of Asian Natural Products Research, 2015
http://dx.doi.org/10.1080/10286020.2015.1041932
C14-Polyacetylene glucosides from Codonopsis pilosula
Yue-Ping Jiang, Yu-Feng Liu, Qing-Lan Guo, Zhi-Bo Jiang, Cheng-Bo Xu,
Cheng-Gen Zhu, Yong-Chun Yang, Sheng Lin and Jian-Gong Shi*
State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of
Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing
100050, China
(Received 29 January 2015; final version received 13 April 2015)
Seven new C14-polyacetylene glucosides codonopilodiynosides A–G (1–7) wereisolated
from an aqueous extract of the Codonopsis pilosula roots. Their structures were
determined by spectroscopic and chemical methods as (–)-(5S,6E,12E)-tetradeca-6,12-
dien-8,10-diyn-1,5,14-triol 5-O-b-D-glucopyranoside (1), (–)-(5S,6E,12E)-tetradeca-
6,12-dien-8,10-diyn-1,5,14-triol 5-O-b-D-glucopyranosyl-(100 ! 20)-b-D-glucopyrano-
side (2), (–)-(5S,6E,12E)-tetradeca-6,12-dien-8,10-diyn-1,5,14-triol 5,14-di-O-b-D-glu-
copyranoside (3), (–)-(5S,6E)-tetradeca-6-en-8,10-diyn-1,5,14-triol 5-O-b-D-
glucopyranoside (4), (–)-(5S,6E,12E)-tetradeca-6,12-dien-8,10-diyn-1,5-diol 5-O-b-D-
glucopyranosyl-(100 ! 20)-b-D-glucopyranoside (5), (–)-(6S,4E,12E)-tetradeca-4,12-
dien-8,10-diyn-1,6-diol 6-O-b-D-glucopyranosyl-(100 ! 20)-b-D-glucopyranoside (6),
and (–)-(5S,6E)-tetradeca-6-en-1,5-epoxy-8,10-diyn-14-ol 14-O-b-D-glucopyranosyl-
(100 ! 20)-b-D-glucopyranoside (7), respectively. The absolute configurations of 1–7
wereassigned by enzymatic hydrolysis followed by isolation of glucoseand aglycones (1a
and4a–7a),andsubsequentcomparisonofspecificrotation,TLC, and1HNMRdataofthe
glucose with an authentic sugar sample and application of modified Mosher’s method
based on the MPA determination rule of Dd RS values for 1a and 4a, and Dd S values for 6a.
The configuration of 7 was assigned byelectronic circular dichroism calculations based on
the quantum-mechanical time-dependent density functional theory.
Keywords: Campanulaceae; Codonopsis pilosula; codonopilodiynosides A–G;
C14-polyacetylene glucosides; absolute configuration
1. Introduction
poor appetite, thirsty, indigestion, chronic
diarrhea, archoptoma, chronic anemia, and
leukemia [1]. Modern pharmacological
studies have revealed that extracts of the
C. pilosula roots have effects in protecting
against peptic ulceration and promoting its
healing, enhancing immunity, and enhan-
cing learning and memory behavior, as well
as inhibiting inducible NO synthase and
protein oxidation and attenuating the
cardiac-impaired insulin-like growth factor
II receptor pathway [2–3]. A variety of
chemical constituents were isolated from
the extracts, such as phytosteroids,
Codonopsis pilosula (Franch.) Nannf. is a
perennial plant of the Campanulaceae
family, which is widely cultivated in the
northwest provinces of China to fulfill
pharmaceutical and food utilizations. The
roots of this plant, known as “dang shen”,
are one of the most common drugs in
traditional Chinese medicine, and some-
times used as a substitute of the much more
costly Panax ginseng as tonic agents
exhibiting similar therapeutic effects [1].
Traditionally C. pilosula roots are mainly
used for the treatment of body weakness,
*Corresponding author. Email: shijg@imm.ac.cn
We dedicate this paper to Prof. Xin-Sheng Yao on the occasion of his 80th birthday.
q 2015 Taylor & Francis

2
Y.-P. Jiang et al.
sesquiterpenes, triterpenes, alkaloids, alkyl
alcohol glycosides, phenylpropanoid gly-
cosides, polyacetylene glycosides, and
polysaccharides [4]. As part of a program
to systematically study the chemical
diversity of traditional Chinese medicines
and their biological effects, focusing on
minor constituents [5–14], an aqueous
decoction of the C. pilosula roots was
investigated since a variety of formulations
containing “dang shen” are practically used
by decocting with water. We report herein
the isolation and structural elucidation of
seven new polyacetylene glucosides (1–7)
(Figure 1) from the decoction.
(c ¼ 0.30, MeOH). Its IR spectrum
showed absorption bands for hydroxyl
(3355 cm21), triple bond (2208 and
2133 cm21), and conjugated double bond
(1565 cm21) functionalities. The molecu-
lar formula of 1, C20H28O8, was indicated
by HR-ESI-MS at m/z 419.1681
[M þ Na]þ combined with NMR spectral
data (Table 1). The 1H NMR spectrum of 1
in DMSO-d6 showed signals attributable to
two trans-disubstituted double bonds at dH
6.48 (dt, J ¼ 15.5 and 4.5 Hz, H-13), 6.27
(dd, J ¼ 15.5 and 5.5 Hz, H-6), 6.08 (d,
J ¼ 15.5 Hz, H-7), and 5.85 (d,
J ¼ 15.5 Hz, H-12); two oxygen-bearing
methylenes at dH 4.05 (d, J ¼ 4.5 Hz, H2-
14), 3.37 (t, J ¼ 5.5 Hz, H2-1); an
oxymethine at dH 4.28 (m, H-5); and
three aliphatic methylenes at dH 1.48 (m,
H2-4), 1.40 (m, H2-2), and 1.31 (m, H2-3),
2. Results and discussion
Compound 1 was obtained as a white
20
amorphous powder with ½aꢀD 268.2
OR2
7
1
3
5
OH
13
11
9
R1O
1
2
3
R1 = H, R2 = β-
D
-glucopyranosyl
-glucopyranosyl-(1"→2')-β-
R1 = H, R2 = β-
D
D
-glucopyranosyl
R1 = R2= β-D-glucopyranosyl
1a R1 = R2= H
OR
OH
HO
4
R = β-
D
-gucopyranosyl
4a R= H
OR
OH
5
R= β-
D-glucopyranosyl-(1"→2')-β-
D-glucopyranosyl
5a R = H
OH
OR
6
R = β-
D
-glucopyranosyl-(1"→2 ')-β-D-glucopyranosyl
6a R = H
1
O
7
3
5
11
9
13
RO
7
R= β-
D-glucopyranosyl-(1"→2 ')-β-
D-glucopyranosyl
7a R = H
Figure 1. The structures of compounds 1–7, 1a, and 4a–7a.

Journal of Asian Natural Products Research
3
d
d
d
d
d
d
d
d
d
d
d
d
d
d
d

4
Y.-P. Jiang et al.
d
d
d
d
d
d
d
d
d
d
d
d
d
d
d

Journal of Asian Natural Products Research
5
in addition to characteristic signals for a b-
glucopyranosyl unit (Table 1). Besides
resonances corresponding to proton-bear-
ing carbons of the above units, the 13C
NMR and DEPT spectra of 1 showed four
quaternary carbon resonances assignable to
two triple bonds (dC 80.8, 80.2, 74.2, and
73.4). These spectral data were consistent
with those of cordifolioidyne B, which was
isolated from C. cordifolioidea [15]. 2D
NMR data analysis confirmed that 1
possessed a planar structure identical to
that of cordifolioidyne B. Especially, in the
HMBC spectrum of 1, correlations from H-
5 to C-10 and from H-10 to C-5 proved that
the b-glucopyranosyl unit was located at
C-5. However, the specific rotation value
of 1 is much smaller than that of
cordifolioidyne B [2151.67 (c ¼ 0.10,
MeOH)]. In addition, the absolute con-
figuration at C-5 in cordifolioidyne B was
not determined in the previous report, and
acid hydrolysis of cordifolioidyne B did not
generate a genuine aglycone instead of a
dehydrated cyclization product of the
aglycone [15]. To determine the absolute
configuration of 1, enzymatic hydrolysis
and modified Mosher’s method based on
the MPA determination rule of Dd RS values
[16] were applied. Hydrolysis of 1 with
snailase produced glucose and 1a. The
glucose was identified by thin-layer liquid
chromatography (TLC) comparison with
an authentic sugar sample, and confirmed
1
by the H NMR spectroscopic data. The
20
positive optical rotation ½aꢀD þ 45.2
(c ¼ 0.06, H2O) of the glucose indicated
20
that it had the same D-configuration as the
authentic
D-glucose
{½aꢀD þ 47.4
1
(c ¼ 0.20, H2O)}. Comparison of the H
NMR and ESI-MS data of 1 and 1a
confirmed that 1a was the aglycone.
Esterification of 1a with (–)-(R)- and
(þ)-(S)-a-methoxyphenylacetic
acid
(MPA) in anhydrous CH2Cl2 [5] afforded
the 1a-tri-(R)- and 1a-tri-(S)-MPA,
respectively. The Dd RS values were calcu-
lated as shown in Figure 2. Application of
Mosher’s rule [16] revealed that 1a had the
5S configuration. Thus, compound 1 was
0.41
H
–0.14
–0.14
–0.06 H
RO–0.06
H H
H
OR
–0.21
H
H
H
H–0.21
H
RO
–0.21
–0.21
0.11
1a-tri-(R)-MPA
1a-tri-(S)-MPA
R = (R)-MPA
R = (S)-MPA
0.40
H
–0.14
–0.09
H –0.09
H
–0.15
R
O
H
H
OR
H
H–0.05
H
H
H –0.26
–0.09
RO
–0.19
0.11
4a-tri-(R)-MPA
4a-tri-(S)-MPA
R = (R)-MPA
R = (S)-MPA
0.26
–0.45
–0H.18
0.2H3
–0.18
H
–0.45
H0.23
H
H
H
OR
H
H
H
0.29
0.08
OR
0.08
6a R = H
6a-bis-(S)-MPA
R = (S)-MPA
Figure 2. Dd RS values (dR –dS, data in blue in ppm) for tri-MPA esters of 1a and 4a, and Dd S
values (dH–dS) for 6a and 6a-bis-(S)-MPA.

6
Y.-P. Jiang et al.
determined as (–)-(5S,6E,12E)-tetradeca-
6,12-dien-8,10-diyn-1,5,14-triol 5-O-b-D-
glucopyranoside and named codonopilo-
diynoside A.
syloxy units locating at different positions
of the aglycone. This was supported by
enzymatic hydrolysis of 3, of which the
hydrolysate was separated to yield 1a and
D-glucose identified by their specific
rotation and 1H NMR spectral data.
In the HMBC spectrum of 3, correlations
from H-5 to C-10, C-3, C-4, C-6, and C-7;
from H-10 to C-5; from H2-14 to C-100, C-
12, and C-13; and from H-100 to C-14
demonstrated that the two b-glucopyrano-
syloxy units were located at C-5 and C-14,
respectively. Therefore, compound 3 was
determined as (–)-(5S,6E,12E)-tetradeca-
6,12-dien-8,10-diyn-1,5,14-triol 5,14-di-
O-b-D-glucopyranoside and named codo-
nopilodiynoside C.
Compound 2, a white amorphous
20
powder with ½aꢀD 222.5 (c ¼ 0.13,
MeOH), has the molecular formula
C26H38O13 as indicated by HR-ESI-MS at
m/z 581.2209 [M þ Na]þ and NMR
spectral data (Table 1). The UV, IR, and
NMR spectroscopic features of 2 were
similar to those of 1, except that the NMR
spectra of 2 exhibited resonances due to an
additional b-glucopyranosyloxy unit repla-
cing one hydroxyl group in 1. Comparison
of the 1H NMR data of 2 and 1
demonstrated that the H-10, H-20, and H-30
and C-20 resonances in the sugar moiety of 2
were deshielded by DdH þ 0.17, þ 0.28,
and þ0.35 ppm and DdC þ 9.0 ppm,
respectively, whereas C-10 and C-30 were
shielded by DdC 21.8 and 20.8 ppm. This
suggested that 2 is an analog of 1 with
substitution of the additional b-D-gluco-
pyranosyl unit at C-20. The suggestion was
further confirmed by enzymatic hydrolysis
and 2D NMR data analysis of 2. By using
the same protocol as described for 1, from
the hydrolysate of 2, aglycone 1a and D-
glucose were isolated and identified. In the
HMBC spectrum, the long-range corre-
lations from H-5 to C-10 and from H-20 to
C-100 confirmed that a b-D-glucopyranosyl-
(100 ! 20)-b-D-glucopyranosyloxy moiety
was located at C-5 in 2. Therefore,
compound 2 was determined as (–)-
(5S,6E,12E)-tetradeca-6,12-dien-8,10-
diyn-1,5,14-triol 5-O-b-D-glucopyranosyl-
(100 ! 20)-b-D-glucopyranoside and named
codonopilodiynoside B.
Compound 4 was obtained as a white
20
amorphous powder with ½aꢀD 234.8
(c ¼ 0.47, MeOH). HR-ESI-MS at m/z
421.1831 [M þ Na]þ indicated that 4 had
the molecular formula C20H30O8, with two
more hydrogen atoms than 1, which was
supported by the NMR spectral data
(Table 1). Comparison of the NMR
spectral data of 4 and 1 indicated the
presence of two sp3 hybridized methylenes
in 4 [dH 2.37 (t, J ¼ 7.2 Hz, H2-12) and
1.68 (2H, quintet, J ¼ 7.2 Hz, H2-13); dC
32.3 (C-13) and 16.6 (C-12)] instead of the
12E double bond in 1. This suggests that 4
is an analog of 1 with saturation of the 12E
double bond. The suggestion was sup-
ported by 2D NMR data analysis of 4,
1
particularly by the H–1H COSY corre-
lations of H2-1/H2-2/H2-3/H2-4/H-5/H-6/
H-7 and H2-12/H2-13/H2-14 and HMBC
correlations from H-5 to C-10, C-4, C-6,
and C-7 and from H2-12 to C-8, C-9, C-10,
C-11, C-13, and C-14. Aglycone 4a and D-
glucose were isolated from enzymatic
hydrolysate of 4, and subsequently ident-
ified by their specific rotation and 1H NMR
spectral data. The 5S-configuration of 4a
was further confirmed by modified
Mosher’s method using Dd RS values of
4a-tri-(R)- and 4a-tri-(S)-MPA (Figure 2).
Thus, compound 4 was determined as (–)-
(5S,6E)-tetradeca-6-en-8,10-diyn-1,5,14-
Compound 3 was obtained as a white
amorphous powder with [a ]2D0 289.8
(c ¼ 0.31, MeOH). The spectroscopic
data of 3 indicated that it is an isomer of
2. Comparing the NMR data of these two
compounds demonstrated that instead of
containing the glucopyranosyl-(100 ! 20)-
b-D-glucopyranosyloxy moiety at C-5 in 2,
compound 3 contains two b-glucopyrano-

Journal of Asian Natural Products Research
7
triol 5-O-b-D-glucopyranoside and named
codonopilodiynoside D.
Compound 5, a white amorphous
(5S,6E,12E)-tetradeca-6,12-dien-8,10-diyn-
1,5-diol 5-O-b-D-glucopyranosyl-(100 ! 20)
-b-D-glucopyranoside and named codono-
pilodiynoside E.
20
powder with ½aꢀD 211.3 (c ¼ 0.12,
MeOH), has the molecular formula
C26H38O12 as indicated by HR-ESI-MS
at m/z 565.2253 [M þ Na]þ. The NMR
spectroscopic data of 5 (Table 1), as
compared with those of 2, indicated that
the terminal hydroxymethylene (CH2-14)
in 2 was replaced by a methyl group [dH
1.76 (dd, J ¼ 6.5 and 1.5 Hz, H3-14; dC
19.2]. 2D NMR data analysis confirmed
that 5 is a 14-deoxygen analog of 2.
Compound 6, a white amorphous
20
powder with ½aꢀD 24.4 (c ¼ 0.03,
MeOH), is an isomer of 5 as indicated by
1
spectroscopic data. Comparison of the H
NMR spectra of 6 and 5 indicated that the
signals of two vicinal methylenes were
shifted from dH 1.40 (H2-3), 1.55 (H-4b),
and 1.66 (H-4a) in 5 to dH 2.05 (H2-3), 2.70
(dd, 17.0 and 5.0 Hz, H-7a), and 2.59 (dd,
17.0 and 7.5 Hz, H-7b), while the double
doublet at dH 6.24 (16.0 and 6.5 Hz, H-6)
and the doublet at dH 5.79 (16.0 Hz, H-7)
for the (6E)-double bond in 5 were replaced
by a double triplet at dH 5.76 (15.5 and
7.0 Hz, H-4) and a double doublet at dH
5.41 (15.5 and 7.5 Hz, H-5) in 6,
respectively. These differences suggested
that the aglycone (5S,6E,12E)-tetradeca-
6,12-dien-8,10-diyn-1,5-diol in 5 is sub-
stituted by a (6S,4E,12E)-tetradeca-4,12-
dien-8,10-diyn-1,6-diol in 6. This was
supported by variation in the 13C NMR
spectral data between the two compounds
(Table 1), as well as confirmed by 2D NMR
data analysis and enzymatic hydrolysis of
6. Particularly, vicinal coupling chain
extensions in 6 were confirmed by the
1H–1H COSY correlations of H2-1/H2-2/
H2-3/H-4/H-5/H-6/H2-7 and H-12/H-13/
H2-14. A diyn-bridged connection between
the coupling chains was confirmed by the
HMBC correlations from H-6 to C-7 and C-
8; from H2-7 to C-5, C-6, C-8, C-9, C-10,
and C-11; and from H3-14 to C-9, C-10, C-
11, C-12, and C-13. In addition, a linkage of
the glucopyranosyl units in 6 was proved by
the HMBC correlations from H-6 to C-10;
from H-10 to C-6; from H-100 to C-20; and
from H-20 to C-100. From the enzymatic
hydrolysate of 6, an aglycone 6a and D-
glucose were isolated, and subsequently
identified by their specific rotation and 1H
NMR spectral data. The 6S-configuration
of 6a was further confirmed by the modified
Mosher’s method using Dd S values of 6 and
1
In particular, the H–1H COSY spectrum
of 5 showed not only a vicinal coupling
correlation between H3-14 and H-13, but
also a long range propenyl coupling
correlation between H3-14 and H-12. The
HMBC spectrum of 5 exhibited two- and
three-bond correlations from H3-14 to C-
13 and C-12, as well as characteristic four-
, five-, and six-bond correlations from H3-
14 to C-11, C-10, and C-9 for long-range
heteronuclear couplings of the enediyne
derivatives, e.g. compounds 1–7 (Sup-
porting Information). In addition, the
HMBC correlations from H-10 to C-5;
from H-5 to C-10, C-3, C-6, and C-7; from
H-100 to C-20; and from H-20 to C-100
confirmed the presence of the b-D-gluco-
pyranosyl-(100 ! 20)-b-D-glucopyranosy-
loxy moiety at C-5 of the aglycone in 5. The
D-configuration of the b-glucopyranosyl
units in 5 was also verified by enzymatic
hydrolysis and subsequent isolation and
identification of the hydrolytic products
using the aforementioned protocol. Agly-
cone 5a isolated from the hydrolysate had
20
an optical rotation value of ½aꢀD þ 15.5
20
(c ¼ 0.07 MeOH) similar to that of 1a
{½aꢀD þ 18.9
(c ¼ 0.05
MeOH)},
suggesting that the C-5 configuration of
5a is identical to that of 1a, though
application of the modified Mosher’s
method failed due to geometric isomeriza-
tion of the 12E double bond in 5a and
limitation of the sample amount. Therefore,
compound 5 was determined as (–)-

8
Y.-P. Jiang et al.
6a-tri-(S)-MPA (Figure 2). Therefore,
compound was determined as
previously undetermined. Because of the
limitation of the sample amount and
instability, the real circular dichroism
(CD) spectrum and specific rotation of 7a
could not be measured. The absolute
configuration of 7 was assigned by
comparing the experimental CD spectrum
of 7 with the calculated electronic circular
dichroism (ECD) spectra of b-D-glucopyr-
anosides of 7a and (5R)-7a predicted from
quantum-mechanical time-dependent den-
sity functional theory (TDDFT) calcu-
lations [18]. The experimental CD of 7 is
in good agreement with that of 7a b-D-
glucopyranoside (Supporting Information
Section). Therefore, compound 7 was
determined as (–)-(5S,6E)-tetradeca-6-en-
1,5-epoxy-8,10-diyn-14-ol 14-O-b-D-glu-
copyranosyl-(100 ! 20)-b-D-glucopyrano-
side and named codonopilodiynoside G.
In the preliminary in vitro assays carried
out in this study, compounds 1, 2, 6, and 7
exhibited weak inhibitory activity against
LPS-induced NO production in BV2-cells
[19], with a maximum inhibition rate of
37.3% at a concentration of 10 mM (the
positive control curcumin gave an inhibition
rate of 86.0% at the same concentration).
These compounds were assessed also for
inhibitory activity against Fe2þ-cysteine-
induced rat liver microsomal lipid peroxi-
dation [20], HIV-1 replication [21], and
several human cancer cell lines [22], but all
were inactive at a concentration of 10 mM.
However, their potential biological activities
and possible contribution to the clinical
effects of this traditional Chinese herbal
medicine are still expected from a further in-
depth evaluation on other biological models,
since polyacetylenes were reported to have
various biological activities, including anti-
bacterial, antifungal, anti-inflammatory, and
anticancer activities [23].
6
(–)-(6S,4E,12E)-tetradeca-4,12-dien-8,10-
diyn-1,6-diol 6-O-b-D-glucopyranosyl-
(100 ! 20)-b-D-glucopyranoside and named
codonopilodiynoside F.
Compound 7 was obtained as a white
20
amorphous powder with ½aꢀD 226.6
(c ¼ 0.04, MeOH). The spectroscopic data
of 7 indicated that it is an isomer of 5. The
NMR spectral data of 7 showed replace-
ment of the 12E double bond and the
methyl group in 5 by two methylenes (dC
16.9 and 29.8) and an oxygen-bearing
methylene (dC 69.3) in 7, respectively.
This, combined with the molecular for-
mula, suggested that the aglycone of 7 was
derived from 5a by saturation of the (12E)-
double bond and hydroxylation of the
methyl, and subsequent dehydration to
form an epoxy ring. The suggestion was
verified by the vicinal coupling correlations
of H2-1/H2-2/H2-3/H2-4/H-5/ H-6/H-7 and
1
H2-12/H2-13/H2-14 in the H–1H COSY
spectrum of 7, together with their chemical
shifts. The diyn-bridged connection
between the coupling chains was confirmed
by the HMBC correlations from H-6 to C-4,
C-5, C-7, and C-8; from H-7 to C-5, C-6, C-
9, and C-10; and from H2-12 to C-8, C-9, C-
10, C-11, C-13 and C-14. In addition, the
HMBC spectrum of 7 showed correlations
from H-10 to C-14; from H2-14 to C-10;
from H-100 to C-20; and from H-20 to C-100,
indicating the presence of the b-D-gluco-
pyranosyl-(100 ! 20)-b-D-glucopyranosyl
moiety at C-14. Accordingly, the epoxy
ring must be formed between the oxygen-
bearing C-1 and C-5 to satisfy with
unsaturation degrees of the molecule,
which was proved by the HMBC corre-
lations from H2-1 to C-5 and from H-5 to C-
1. Aglycone 7a and D-glucose were isolated
from the enzymatic hydrolysate of 7 and
identified by using the same protocol as
described above. Although a natural
product having the identical planar struc-
ture with 7a was reported from Campanula
glomerata [17], its configuration was
3. Experimental
3.1 General experimental procedures
Optical rotations were measured on P-2000
polarimeter (JASCO, Tokyo, Japan). UV

Journal of Asian Natural Products Research
spectra were measured on a V-650 3.2 Plant material
9
spectrometer (JASCO, Tokyo, Japan). IR
spectra were recorded on a Nicolet 5700
FT-IR microscope instrument (FT-IR
microscope transmission) (Thermo Elec-
tron Corporation, Madison, WI, USA).
NMR spectra were obtained at 500 MHz or
600 MHz for 1H, and 125 MHz or 150 MHz
for 13C, respectively, on Inova 500 or SYS
600 (Varian Associates Inc., Palo Alto, CA,
USA) or Bruker 600 NMR spectrometers
(Bruker Corp., Karlsruhe, Germany) in
DMSO-d6, MeOH-d4, or acetone-d6, with
solvent peaks used as references. ESI-MS
and HR-ESI-MS data were measured using
an AccuToFCS JMS-T100CS spec-
trometer (Agilent Technologies, Ltd.,
Santa Clara, CA, USA). Column chroma-
tography (CC) was performed with silica
gel (200–300 mesh, Qingdao Marine
Chemical Inc. Qingdao, China), Sephadex
LH-20 (Pharmacia Biotech AB, Uppsala,
Sweden), CHP 20P (Mitsubishi Chemical
Inc., Tokyo, Japan), or HW-40F (Tosoh
Bioscience LLC., Tokyo, Japan). HPLC
separation was performed on an instrument
consisting of an Agilent ChemStation for a
LC system, an Agilent1200 pump, and an
Agilent 1100 single-wavelength absor-
bance detector (Agilent Technologies,
Ltd.) with a YMC-Pack Ph (250 £ 10 mm
i.d.) column packed with phenyl-silica gel
(5 mm) (YMC Co., Ltd, Kyoto, Japan), a
Spherigel (300 £ 10 mm) semipreparative
column packed with C8 reversed phase
silica gel (5 mm) (Johnson Inc., Dalian,
China) or a Grace (250 £ 10 mm) semi-
preparative column packed with C18
reversed phase silica gel (5 mm) (W.R
Grace & Co., Maryland, USA). TLC was
carried out with glass precoated silica gel
GF254 plates (Qingdao Marine Chemical
Inc., Qingdao, China). Spots were visual-
ized under UV light or by spraying with 7%
H2SO4 in 95% EtOH followed by heating.
Unless otherwise noted, all chemicals were
obtained from commercially available
sources and were used without further
purification.
The roots of Codonopsis pilosula (Franch.)
Nannf. were collected in October 2012
from the culture field in Weiyuan, Gansu
Province, China. Plant identity was ver-
ified by Mr Lin Ma (Institute of Materia
Medica, Beijing 100050, China).
A voucher specimen (no. ID-S-2503) was
deposited at the herbarium of the Depart-
ment of Natural Medicinal Chemistry,
Institute of Materia Medica, Beijing
100050, China.
3.3 Extraction and isolation
The dried and powered roots of C. pilosula
(50 kg) were decocted with H2O (150 l,
3 £ 0.5 h). The decoction was evaporated
under reduced pressure to yield a brown
residue (26 kg). The residue was dissolved
in H2O (100 l), loaded on a macroporous
adsorbent resin (HPD-110, 20 l) column
(20 £ 200 cm), and eluted successively
with H2O (100 l), 50% EtOH (120 l), and
95% EtOH (80 l) to yield three correspond-
ing fractions A–C. Fraction B (270 g) was
chromatographed over MCI gel CHP 20P
(5.5 L), successively eluting with H2O
(20 l), 30% EtOH (30 l), 50% EtOH (30 l),
and 95% EtOH (8 l), to give B1–B4.
Fraction B3 (22 g) was subjected to flash
chromatography over RP silica gel, eluting
with a gradient of increasing MeOH (40–
100%) in H2O, to yield subfractions B3-1–
B3-5. B3-1 (10 g) was fractionated by CC
over silica gel, eluting with a gradient of
increasing MeOH (0–100%) in CHCl3, to
yield B3-1-1–B3-1-10, of which B3-1-9
(1.1 g) was further separated by CC over
HW-40F using MeOH as the mobile phase
to give B3-1-9-1–B3-1-9-5. Fraction B3-
1-9-1 (0.3 g) was isolated by preparative
silica gel TLC (EtOAc–n-butanol, 1:2) to
yield B3-1-9-1-1–B3-1-9-1-3. Purification
of B3-1-9-1-1 (75 mg) by RP-HPLC (C8
column, 1.5 ml/min, UV 205 nm) with
CH3CNZH2O (28:72) as the mobile
phase yielded 2 (5.0 mg, tR ¼ 16 min),
while B3-1-9-1-2 (150 mg) and B3-1-9-1-

10
Y.-P. Jiang et al.
3 (60 mg) were separately purified by RP
HPLC (C18 column, 1.5 ml/min, UV
205 nm) using the mobile phase of 50%
MeOH in H2O for the former to afford 1
(6.8 mg, tR ¼ 26 min) and 47% MeOH in
H2O for the later to give 4 (7.0 mg,
tR ¼ 41 min). Fraction B3-1-10 (1.0 g)
was separated by CC over HW-40F
(MeOH) to give B3-1-10-1–B3-1-10-6, of
which B3-1-10-2 (100 mg) was purified by
RP HPLC (C18 column, 1.5 ml/min, UV
220 nm, 17% CH3CN in H2O) to yield 3
(4.0 mg, tR ¼ 30 min). Fractionation of B3-
2 (5.3 g) by CC over silica gel, eluting with
a gradient of increasing MeOH (0–100%)
in CHCl3, yielded B3-2-1–B3-2-6, and B3-
2-1(1.0 g) was further fractionated by CC
over Sephadex LH-20 (CHCl3 2
MeOH, 1:1) to give B3-2-1-1–B3-2-1-8.
Purification of B3-2-1-1 (90 mg) by RP
HPLC (C18 column, 1.5 ml/min, UV
220 nm, 50% MeOH in H2O) afforded 6
(4.2 mg, tR ¼ 61 min). Separation of B3-3
(1.5 g) by RP HPLC (C18 column, 3.0 ml/
min, UV 220 nm, 29% CH3CN in H2O)
yielded B3-3-1–B3-3-6, of which B3-3-4
(60 mg) was purified by RP HPLC (C8
column, 1.5 ml/min, UV 220 nm, 29%
CH3CN in H2O) to afford 5 (3.9 mg,
tR ¼ 56 min), and B3-3-5 (80 mg) by RP
HPLC (C18 column, 1.5 ml/min, UV
205 nm 26% CH3CN in H2O) to yield 7
(5.0 mg, tR ¼ 67 min).
3.3.2 Codonopilodiynoside B (2)
20
White amorphous powder; ½aꢀD 222.5
(c 0.13, MeOH); UV (MeOH) lmax (log e)
216 (4.29), 230 (4.29), 237 (4.32), 247
(4.20 sh), 262 (3.78), 277 (4.00), 294
(4.17), 314 (4.09) nm; IR nmax 3378, 2924,
2208, 2133, 1647, 1601, 1415, 1370, 1079,
898, 629 cm21 1H NMR (DMSO-d6,
;
600 MHz) spectral data, see Table 1; 13C
NMR (DMSO-d6, 150 MHz) spectral data,
see Table 1; (þ)-ESI-MS m/z 581
[M þ Na]þ, 597 [M þ K]þ; (2)-ESI-
MS: m/z 557 [M 2 H]2, 593 [M þ Cl]2;
HR-ESI-MS: m/z 581.2209 [M þ Na]þ
(calcd for C26H38O13Na, 581.2205).
3.3.3 Codonopilodiynoside C (3)
20
White amorphous powder; ½aꢀD 289.8
(c 0.31, MeOH); UV (MeOH) lmax (log e)
203 (5.18), 263 (4.36), 278 (4.60), 295
(4.73), 314 (4.65) nm; IR nmax 3365, 2927,
2878, 2206, 2129, 1637, 1511, 1414, 1364,
1315, 1268, 1158, 1077, 1041, 958, 898,
1
629 cm21; H NMR (CD3OD, 500 MHz)
spectral data, see Table 1; 13C NMR
(CD3OD, 125 MHz) spectral data, see
Table 1; (þ)-ESI-MS: m/z 581
[M þ Na]þ, 597 [M þ K]þ; (–)-ESI-MS:
m/z 557 [M 2 H]2, 593 [M þ Cl]2; HR-
ESI-MS: m/z 581.2204 [M þ Na]þ (calcd
for C26H38O13Na, 581.2205).
3.3.1 Codonopilodiynoside A (1)
20
3.3.4 Codonopilodiynoside D (4)
20
White amorphous powder; ½aꢀD 268.2
White amorphous powder; ½aꢀD 234.8
(c 0.30, MeOH); UV (MeOH) lmax (log e)
215 (3.31), 236 (3.32), 263 (2.64), 278
(2.95), 294 (3.14), 314 (3.05) nm; IR nmax
3355, 2927, 2870, 2208, 2133, 1565, 1413,
(c 0.47, MeOH); UV (MeOH) lmax (log e)
206 (4.06, sh), 212 (4.10), 240 (3.26), 253
(3.49), 268 (3.66), 284 (3.56) nm; IR nmax
3395, 3188, 3011, 2922, 2850, 2236, 1646,
1468, 1420, 1119, 1076, 1044, 960, 816,
1371, 1077, 1042, 957, 659 cm21 1H
;
NMR (DMSO-d6, 500 MHz) spectral data,
see Table 1; 13C NMR (DMSO-d6,
125 MHz) spectral data, see Table 1;
(þ)-ESI-MS: m/z 419 [M þ Na]þ; (2)-
ESI-MS m/z 431 [M þ Cl]2; HR-ESI-MS:
m/z 419.1681 [M þ Na]þ (calcd for
C20H28O8Na, 419.1676).
722, 647 cm21 1H NMR (CD3OD,
;
600 MHz) spectral data, see Table 1; 13C
NMR (CD3OD, 150 MHz) spectral data,
see Table 1; (þ)-ESI-MS: m/z 421 [M þ
Na]þ; (2)-ESI-MS: m/z 433 [M þ Cl]2;
HR-ESI-MS: m/z 421.1831 [M þ Na]þ
(calcd for C20H30O8Na, 421.1833).

Journal of Asian Natural Products Research
11
3.3.5 Codonopilodiynoside E (5)
20
[M þ Na]þ (calcd for C26H38O12Na,
565.2255).
White amorphous powder; ½aꢀD 211.3
(c 0.12, MeOH); UV (MeOH) lmax (log e)
203 (4.48), 263 (3.81), 277 (4.05), 294
(4.20), 313 (4.11) nm; IR nmax 3390, 3012,
2922, 2850, 2206, 2130, 1646, 1468, 1419,
1365, 1076, 1301, 1271, 1166, 1077, 1038,
3.3.8 Enzymatic hydrolysis of 1–7
Compounds 1 2 7 (3.0–5.0 mg) were
separately hydrolyzed in H2O (3 ml) with
snailase at 378C for 24 h, then extracted by
EtOAc (3 £ 3 ml). The aqueous phase was
dried by a stream of N2, then chromato-
graphed over silica gel eluting with
CH3CNZH2O (8:1), to yield glucose.
The glucose (0.75–2.47 mg) from the
hydrolysates of 1 2 7 gave retention
factor (Rf < 0.33; CH3CNZH2O, 6:1) on
954, 896, 721, 643 cm21
;
1H NMR
(CD3OD, 500 MHz) spectral data, see
Table 1; 13C NMR (CD3OD, 125 MHz)
spectral data, see Table 1; (þ)-ESI-MS: m/
z 565 [M þ Na]þ; (2)-ESI-MS: m/z 541
[M 2 H]2, 577 [M þ Cl]2; HR-ESI-MS:
m/z 565.2253 [M þ Na]þ (calcd for
C26H38O12Na, 565.2255).
20
TLC, ½aꢀD values [þ43.1 to þ48.1
(c ¼ 0.06–0.19, H2O)], and 1H NMR
spectral data (D2O) consistent with those
of an authentic D-glucose. The EtOAc
phase was concentrated under reduced
pressure, and purified by RP-HPLC to
3.3.6 Codonopilodiynoside F (6)
20
Amorphous white powder; ½aꢀD 24.4
(c 0.03, MeOH); UV (MeOH) lmax (log e)
212 (3.98), 240 (2.58), 252 (3.19), 266
(3.39), 282 (3.26) nm; IR nmax 3392, 2924,
2234, 2143, 1647, 1421, 1367, 1304, 1265,
1166, 1077, 1037, 953, 898, 634 cm21; 1H
NMR (DMSO-d6, 500 MHz) spectral data,
see Table 1; 13C NMR (DMSO-d6,
125 MHz) spectral data, see Table 1;
(þ)-ESI-MS: m/z 565 [M þ Na]þ, 581
[M þ K]þ; (2)-ESI-MS: m/z 541
[M 2 H]2, 577 [M þ Cl]2; HR-ESI-MS:
m/z 565.2253 [M þ Na]þ (calcd for
C26H38O12Na, 565.2255).
20
obtain 1a (0.30–1.00 mg) {½aꢀD þ 12.6 to
20
þ15.5 (c ¼ 0.02–0.08, MeOH)} from
1–3, 4a {1.09 mg, ½aꢀD þ 29.5
20
(c ¼ 0.08, MeOH)}, 5a {0.65 mg,
20
½aꢀD þ 18.9 (c ¼ 0.05, MeOH)}, 6a
{0.52 mg, ½aꢀD þ 10.0 (c ¼ 0.04,
MeOH)}, and 7a (0.78 mg) from 4–7,
respectively. Compound 1a: 1H NMR
(600 MHz, acetone-d6)
d 6.46 (dt,
J ¼ 15.6, 4.2 Hz, H-13), 6.40 (dd,
J ¼ 15.6, 5.4 Hz, H-6), 5.89 (dt,
J ¼ 15.6, 1.8 Hz, H-12), 5.87 (d,
J ¼ 15.6 Hz, H-7), 4.19 (m, H-5 and
H-14), 4.11 (t, J ¼ 5.4 Hz, OH-14), 4.08
(d, J ¼ 4.8 Hz, OH-5), 3.53 (t, J ¼ 6.0 Hz,
H-1), 3.40 (t, J ¼ 5.4 Hz, OH-1), 1.52 (m,
H-2, H-3, and H-4a), 1.43 (m, H-4b).
Compound 4a: 1H NMR (600 MHz,
MeOH-d4) d 6.22 (dd, J ¼ 16.2, 6.0 Hz,
H-6), 5.69 (d, J ¼ 16.2 Hz, H-7), 4.07 (q,
J ¼ 6.0 Hz, H-5), 3.59 (t, J ¼ 6.0 Hz,
H-1), 3.52 (t, J ¼ 6.6 Hz, H-14), 2.39 (t,
J ¼ 6.6 Hz, H-12), 1.70 (quintet,
J ¼ 6.6 Hz, H-13), 1.49 (m, H-2 and 3),
1.44 (m, H-4a), 1.37 (m, H-4b). Com-
pound 5a: 1H NMR (600 MHz, acetone-d6)
d 6.39 (dd, J ¼ 15.6, 5.4 Hz, H-6), 6.36
(dq, J ¼ 15.6, 7.2 Hz, H-13), 5.86 (dd,
3.3.7 Codonopilodiynoside G (7)
20
White amorphous powder; ½aꢀD 226.6
(c 0.04, MeOH); UV (MeOH) lmax (log e)
206 (3.71), 240 (3.04), 253 (3.24), 268
(3.39), 283 (3.33) nm; IR nmax 3395, 2922,
2850, 2230, 1646, 1468, 1420, 1077, 1041,
895, 645 cm21
;
1H NMR (CD3OD þ
DMSO-d6, 600 MHz) spectral data, see
Table 1; 13C NMR (CD3OD þ DMSO-d6,
150 MHz) spectral data, see Table 1; (þ)-
ESI-MS: m/z 565 [M þ Na]þ, 581
[M þ K]þ; (2)-ESI-MS: m/z 541
[M 2 H]2, 577 [M þ Cl]2; HR-ESI-MS:
m/z 543.2435 [M þ H]þ (calcd for
C26H39O12,
543.2436),
565.2261

12
Y.-P. Jiang et al.
J ¼ 15.6 Hz, H-7), 5.68 (dq, J ¼ 15.6,
1.8 Hz, H-12), 4.35 (d, J ¼ 5.4 Hz, OH-5),
4.20 (m, H-5), 3.70 (t, J ¼ 5.4 Hz, OH-1),
3.54 (m, H-1), 1.83 (dd, J ¼ 7.2, 1.8 Hz,
H-14), 1.52 (m, H-2,3, and 4a), 1.43 (m, H-
(6H, s, 2 £ MeO), 3.37 (3H, s, MeO), 1.50
(2H, m, H-3), 1.39 (4H, m, H-2,4). 1a-tri-
(S)-MPA: 1H NMR (600 MHz, acetone-d6)
d 7.34 2 7.46 (15H, m, aromatic H), 6.36
(1H, dt, J ¼ 16.2, 5.4 Hz, H-13), 6.18(1H,
dd, J ¼ 16.2, 6.0 Hz, H-6), 5.76 (1H, d,
J ¼ 16.2 Hz, H-12), 5.42 (1H, d,
J ¼ 16.2 Hz, H-7), 5.32 (1H, m, H-5),
4.92 (1H, s, a-H), 4.89 (1H, s, a-H), 4.83
(1H, s, a-H), 4.73 (2H, dd, J ¼ 5.4, 1.8 Hz,
H-14), 4.06 (2H, dt, J ¼ 6.0, 1.8 Hz, H-1),
1.60 (4H, m, H-2,4), 1.56 (2H, m, H-3).
4a-tri-(R)-MPA: 1H NMR (500 MHz,
acetone-d6) d 7.34 2 7.45 (15H, m, aro-
matic H), 6.23 (1H, dd, J ¼ 16.0, 6.0 Hz,
H-6), 5.76 (1H, d, J ¼ 16.0 Hz, H-7), 5.27
(1H, m, H-5), 4.90 (1H, s, a-H), 4.87 (1H,
s, a-H), 4.80 (1H, s, a-H), 4.18 (2H, t,
J ¼ 5.5 Hz, H-14), 3.91 (2H, t, J ¼ 6.0 Hz,
H-1), 3.38 (6H, s, MeO), 3.36 (3H, s,
MeO), 2.35 (2H, t, J ¼ 7.0 Hz, H-12), 1.83
(2H, m, H-13), 1.49 (2H, m, H-3), 1.39
(3H, m, H-2, H-4a), 0.99 (1H, m, H-4b).
4a-tri-(S)-MPA: 1H NMR (500 MHz,
acetone-d6) d 7.34 2 7.45 (15H, m, aro-
matic H), 6.12 (1H, dd, J ¼ 16.0, 5.5 Hz,
H-6), 5.36 (1H, d, J ¼ 16.0 Hz, H-7), 5.30
(1H, m, H-5), 4.88 (1H, s, a-H), 4.86 (1H,
s, a-H), 4.83 (1H, s, a-H), 4.17 (2H, t,
J ¼ 5.0 Hz, H-14), 4.06 (2H, t, J ¼ 6.0 Hz,
H-1), 3.38 (9H, s, MeO), 2.34 (2H, t,
J ¼ 7.5 Hz, H-12), 1.83 (2H, m, H-13),
1.58 (5H, m, H-2, 3, 4a), 1.25 (1H, m, H-
4b). 6a-bis-(S)-MPA: 1H NMR (600 MHz,
acetone-d6) d 7.31 2 7.47 (10H, m, aro-
matic H), 6.35 (1H, dq, J ¼ 15.6, 6.6 Hz,
H-13), 5.62 (1H, d, J ¼ 15.6 Hz, H-12),
5.45 (1H, dt, J ¼ 15.6, 7.2 Hz, H-4), 5.35
(1H, m, H-6), 5.31 (1H, m, H-5), 4.86 (1H,
s, a-H), 4.84(1H, s, a-H), 4.00 (2H, m, H-
1), 3.39 (3H, s, MeO), 3.38 (3H, s, MeO),
2.69 (2H, m, H-7), 1.87 (q, J ¼ 7.2 Hz, H-
3), 1.81 (3H, dd, J ¼ 6.6, 1.8 Hz, H-14),
1.52 (2H, m, H-2).
1
4b). Compound 6a: H NMR (600 MHz,
acetone-d6) d 6.30 (dq, J ¼ 15.6, 6.6 Hz,
H-13), 5.74 (dt, J ¼ 15.6, 6.6 Hz, H-4),
5.59 (dq, J ¼ 15.6, 1.8 Hz, H-12), 5.57
(dd, J ¼ 15.6, 4.8 Hz, H-5), 4.21 (m, H-6),
4.13 (d, J ¼ 5.4 Hz, OH-6), 3.55 (m, H-1),
3.42 (t, J ¼ 5.4 Hz, OH-1), 2.51 (m, H-7),
2.10 (q, J ¼ 7.2 Hz, H-3), 1.79 (dd,
J ¼ 6.6, 1.8 Hz, H-14), 1.58 (quintet,
J ¼ 7.2 Hz, H-2). Compound 7a: 1H
NMR (600 MHz, MeOH-d4) d 6.26 (dd,
J ¼ 15.6, 5.4 Hz, H-6), 5.74 (d,
J ¼ 15.6 Hz, H-7), 3.99 (m, H-5), 3.94
(br d, J ¼ 12.0 Hz, H-1a), 3.88 (m, H-1b),
3.61 (t, J ¼ 6.0 Hz, H-14), 2.44 (t,
J ¼ 6.0 Hz, H-12), 1.81 (m, H-3a), 1.72
(quintet, J ¼ 6.0 Hz, H-13), 1.69 (m, H-
3b), 1.48 2 1.58 (m, H-2 and 4).
3.3.9 Synthesis of (R)- and (S)-MPA
esters of 1a, 4a, and 6a
To a solution of 1a, 4a, or 6a (0.3–0.5 mg)
in anhydrous CH2Cl2 (3.0 ml), (R)-MPA or
(S)-MPA (5.3 mg), 1-ethyl-3-(3-dimethyl-
laminopropyl)carbodiimide hydrochloride
(6.1 mg), and 4-dimethylaminopyridine
(3.9 mg) were added, then kept at rt for 15
min [5]. After evaporation of the solvent
under reduced pressure, the residue was
separated by preparative TLC (petroleum
ether-acetone, 3:1) to yield 1a-tri-(R)- and
(S)-MPA from 1a, 4a-tri-(R)- and (S)-
MPA from 4a, and 6a-bis-(S)- from 6a,
respectively. 1a-tri-(R)-MPA: 1H NMR
(600 MHz, acetone-d6) d 7.32 2 7.46
(15H, m, aromatic H), 6.37 (1H, dt,
J ¼ 16.2, 5.4 Hz, H-13), 6.29 (1H, dd,
J ¼ 16.2, 6.0 Hz, H-6), 5.83 (1H, d,
J ¼ 16.2 Hz, H-7), 5.77 (1H, d,
J ¼ 16.2 Hz, H-12), 5.29 (1H, m, H-5),
4.92 (1H, s, a-H), 4.91 (1H, s, a-H), 4.80
(1H, s, a-H), 4.73 (2H, dd, J ¼ 5.4, 1.8 Hz,
H-14), 3.92 (2H, t, J ¼ 6.0 Hz, H-1), 3.39
3.3.10 ECD calculation
Conformational analysis of b-D-glucopyr-
anosides of 7a and (5R)-7a was performed

Journal of Asian Natural Products Research
by using the MMFF94 molecular mech- References
13
anics force field via the Molecular
Operating Environment software package
[24]. Each molecule showed one lowest
energy conformer whose relative energy
was within 2 kcal/mol (Supporting Infor-
mation Section). The conformer was
further optimized at the B3LYP/6-31g(d,
p) level in MeOH. The energies, oscillator
strengths, and rotational strengths of the
first 30 electronic excitations were calcu-
lated using the TDDFT methodology at the
B3LYP/6-311þþG (2d, 2p) level. ECD
spectrum of the conformer was simulated
using a Gaussian function with a half-
bandwidth of 0.28 eV. The corresponding
theoretical ECD spectrum of the enantio-
mer was depicted by inverting that of the
model compound. All quantum compu-
tations were performed using Gaussian 09
program package [25], on an IBM cluster
machine located at the High Performance
Computing Center of Peking Union
Medical College. In the 200–400 nm
region, the theoretically calculated ECD
spectra of 7a b-D-glucopyranoside was in
agreement with the experimental ECD
spectra of 7 (Supporting Information).
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Disclosure statement
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No potential conflict of interest was reported
by the authors.
Funding
Financial support from the National Natural
Sciences Foundation of China [grant number
30825044], [grant number 21132009]; the
Program for Changjiang Scholars and Innova-
tive Research Team in University [grant
number IRT1007]; and the National Science
and Technology Project of China [grant number
2012ZX09301002-002], [grant number
2011ZX0 9307-002-01] is acknowledged.
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