Hypoglycemic triterpenoid glycosides from Cyclocarya paliurus (Sweet Tea Tree)

Article abstract of DOI:10.1016/j.bioorg.2019.103493

Four new rarely occurred seco-dammarane triterpenoid glycosides (1–4) and four new dammarane triterpenoid glycosides (5–8), along with four known triterpenoids (9–12), were isolated from the 70% ethanol extract of the leaves of Cyclocarya paliurus (family Juglandaceae). Their structures were elucidated by extensive spectroscopic methods, including 1D/2D NMR and HRESIMS data, together with chemical analysis and DFT GIAO ¹³C NMR calculation. In bioassay, compounds 5–8 significantly increased glucose consumption in 3T3-L1 adipocytes, which could be the bioactive constituents for the anti-diabetes effect of the traditional usage of C. paliurus.

Full text of DOI:10.1016/j.bioorg.2019.103493

Journal Pre-proofs
Hypoglycemic Triterpenoid Glycosides from Cyclocarya paliurus (Sweet Tea
Tree)
Huihui Sun, Jie Tan, Wenyan Lv, Jing Li, Jianping Wu, Jialing Xu, Hui Zhu,
Zhichun Yang, Wenxuan Wang, Zijun Ye, Tongyao Xuan, Zhenxing Zou,
Zuhui Chen, Kangping Xu
PII:
S0045-2068(19)31824-3
DOI:
https://doi.org/10.1016/j.bioorg.2019.103493
YBIOO 103493
Reference:
To appear in:
Bioorganic Chemistry
Received Date:
Revised Date:
Accepted Date:
28 October 2019
26 November 2019
1 December 2019
Please cite this article as: H. Sun, J. Tan, W. Lv, J. Li, J. Wu, J. Xu, H. Zhu, Z. Yang, W. Wang, Z. Ye, T. Xuan,
Z. Zou, Z. Chen, K. Xu, Hypoglycemic Triterpenoid Glycosides from Cyclocarya paliurus (Sweet Tea Tree),
Bioorganic Chemistry (2019), doi: https://doi.org/10.1016/j.bioorg.2019.103493
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Hypoglycemic Triterpenoid Glycosides from Cyclocarya paliurus (Sweet Tea Tree)
Huihui Suna, Jie Tana, Wenyan Lva, Jing Lia, Jianping Wua, Jialing Xua, Hui Zhua, Zhichun Yanga,
Wenxuan Wanga, Zijun Yeb, Tongyao Xuanb, Zhenxing Zoua, Zuhui Chenc, Kangping Xua,*
aXiangya School of Pharmaceutical Sciences, Central South University, Changsha 410013, PR China
bCentral South University Xiangya School of Medicine Affiliated HaiKou Hospital, Haikou 570208,
PR China
cHunan Heran Biotechnology Development company, Changsha 410000, PR China
*Corresponding author.
E-mail address: xukp395@csu.edu.cn (Kangping Xu)

Abstract
Four new rarely occurred seco-dammarane triterpenoid glycosides (1-4) and four new
dammarane triterpenoid glycosides (5-8), along with four known triterpenoids (9-12), were isolated
from the 70% ethanol extract of the leaves of Cyclocarya paliurus (family Juglandaceae). Their
structures were elucidated by extensive spectroscopic methods, including 1D/2D NMR and
13
HRESIMS data, together with chemical analysis and DFT GIAO C NMR calculation. In bioassay,
compounds 5-8 significantly increased glucose consumption in 3T3-L1 adipocytes, which could be
the bioactive constituents for the anti-diabetes effect of the traditional usage of C. paliurus.
Keywords: Cyclocarya paliurus; triterpenoid glycosides; glucose consumption; 3T3-L1 adipocytes

1. Introduction
Cyclocarya paliurus (Batal.) Iljinsk (family Juglandaceae), an endemic species in China, is
widely distributed in south and southeast areas [1]. Its leaves are used as traditional tea, which have
the effects of relieving diabetes, hypertension, coronary heart disease and neurasthenia [2, 3]. To date,
various compounds including triterpenoids [4], flavonoids [5], phenolic acids [5], polysaccharides [6]
and steroids [7] have been isolated from this plant. Especially, triterpenoids from C. paliurus have
been the focus of research due to their interesting pharmacological activity on diabetes [8-13].
During our previous study, the dichloromethane, ethyl acetate and n-butanol portions of the total
70% ethanol extract of C. paliurus were evaluated for their effects on body weight, water intake,
fasting blood glucose and fasting insulin values of type-II diabetic rats. As a result, dichloromethane
portion showed significant effects and alleviated the insulin resistance symptoms of type-II diabetic
rats. Further investigation on the dichloromethane portion led to the isolation of triterpenoid
glycosides 1-12 (Fig. 1). Herein, their isolation, structure elucidation, and hypoglycemic activity
assay were reported.

HO
HO
OH
24
27
26
OH
OH
OH
OH
OH
OH
O
OH
20
21
H
O
O
5'
23
H
HO
HO
HO
HO
O
O
O
1'
H
O
H
HO
O
OH
OH
H
H
O
OH
12
3'
1
17
H
OH
18
OH
O
O
H
O
19
H
13
H
9
O
HO
H3CO
3
H
HO
15
H
H
H
30
28
HO
4
H
6
29
4*
2*
1*
3
*
OH
OH
OH
5'
OH
OH
20
O
O
HO
HO
O
1'
O
O
O
H
O
O
HO
H
HO
HO
O
12
HO
HO
O
3'
1
H
OH
H
OH
18
H
H
OH
19
OH
9
14
16
H
H
H
4
H
OH
HO
OH
3
30
H
H
7
H
O
O
5
5''
O
H
O
4''
HO
O
O
O
O
1''
HO
HO
28
29
HO
HO
OH
OH
OH
OH
8*
5*
7*
6*
OH
OH
HO
OH
OH
OH
HO
OH
OH
OH
OH
O
O
O
O
OH
O
O
O
O
H
HO
O
HO
O
OCH3
OH
OH
H
H
H
H
OH
OH
O
O
H
H
H
HO
HO
HO
HO
H
H
H
H
11
12
10
9
Fig. 1. Chemical structures of compounds 1-12.

2. Materials and methods
2.1. General methods
Optical rotations were measured on a JASCO model 1020 polarimeter (Horiba, Tokyo, Japan) at
room temperature. NMR spectra were measured on a Bruker AV-500 MHz spectrometer (Bruker,
Karlsruhe, Germany) using acetone-d6 or CD3OD as solvent and tetramethylsilane (TMS) as an
internal standard. The HRESIMS were measured on a LC-LTQ Orbitrap Velos Pro ETD (Thermo
Fisher, MA, USA) in positive ion mode. Gas Chromatography-Mass Spectrometer (GC-MS) were
measured on GCMS-QP2010 Ultra (SHIMADZU, Hongkong, China). Column chromatograph (CC)
separations were carried out on silica gel (200-300 or 80-100 mesh; Qingdao Peremanent Sea Silica
Ltd., Qingdao, China), polyamide (80-100 or 30-60 mesh, Taizhou Luqiao Sijia Biochemical Plastics
Factory, Taizhou, China), MCI gel (75-150 µm, Mitsubishi Chemical Industries Ltd., Tokyo, Japan),
C18 reverse-phased silica gel (40-75 µm, Fuji, Kasugai, Japan). Semi-preparative HPLC separations
were conducted by using Agilent 1260 system with an YMC-peak ODS-A column (5 µm, 250 × 10
mm). All the analytic reagents were analytical grade and purchased from Cologne Chemical Co., Ltd.,
Chengdu, China.
2.2. Plant material
The leaves of C. paliurus (collected in Xinning County, Shaoyang City, Hunan Province) were
provided by Hunan Heran Biotechnology Development Company, Hunan Province, People's Republic
of China, in May 2016, identified by Prof. Kangping Xu (College of Xiangya Pharmacy, Central

South University). And its voucher specimen (No. 20160820) was deposited in the Xiangya School of
Pharmaceutical Sciences, Central South University.
2.3. Extraction and isolation
The air-dried leaves of C. paliurus (10.0 kg) were extracted with 70% EtOH under reflux (100 L;
2 × 2 h) to get crude extracts, which were dried under reduced pressure afterwards. Then the residue
was dispersed in water and partitioned with with CH2Cl2, EtOAc and n-BuOH, sequentially. The
CH2Cl2 extract (400 g) was subjected to column chromatography on silica gel and eluted with a
gradient mixture of CH2Cl2/MeOH (from 100:0 to 0:100) to obtain ten fractions (I-X) according to
TLC patterns. The fraction IX was eluted with aqueous through Polyamide column (0-95%, stepwise)
to yield five subfractions (A-E). IXA (33.2 g) was further chromatographed by a silica gel column
(CH2Cl2-MeOH, from 50:1 to 0:100), and followed by a C18 reversed-phase column (from 20% to
100% aqueous MeOH, stepwise) to obtain compound 1 (2.3 mg) and compound 2 (4.5 mg).
Fr.IXB-Fr.IXC were merged together and then separated by a C18 reversed-phase column with a
gradient system of MeOH/H2O to get five fractions (IXBC-1-IXBC-5). IXBC-2 (3.5 g) was purified by an
MCI gel column eluted with MeOH-H2O system (20% to 100%, stepwise) to give six subfractions
(IXBC-2-a-IXBC-2-f). Compound 3 (2.0 mg), compound 4 (3.2 mg) were isolated from IXBC-2-b by
semi-preparative HPLC (3.0 mL/min, 210 nm, ACN-H2O, 7.0:3.0, V/V). Compound 6 (1.8 mg),
compound 7 (1.5 mg) were isolated from IXBC-2-c by semi-preparative HPLC (3.0 mL/min, 230 nm,
ACN-H2O, 7.0:3.0, V/V). Fraction IXD (78.2 g) was applied to silica gel eluted with EtOAc-MeOH to
give eight fractions (IXD-1-IXD-8). The fraction IXD-3 (18.4 g) was futher purified by silica gel eluted
with CH2Cl2-MeOH to give four fractions (IXD-3-a-IXD-3-d). Subsequently, Compound 5 (3.8 mg) was

isolated from IXD-3-b by semi-preparative HPLC (3.0 mL/min, 230 nm, ACN-H2O, 8.0:2.0, V/V).
Compound 8 (2.5 mg) and compound 9 (4.8mg) were isolated from IXD-3-c by semi-preparative HPLC
(3.0 mL/min, 230 nm, ACN-H2O, 8.0:2.0, V/V). Next, IXD-4 (23.0 g) was purified by an MCI gel
column eluted with aqueous MeOH (20% to 100%, stepwise) to give six subfractions (IXD-4-a-IXD-4-e).
Compound 10 (8.4 mg) was isolated from IXD-4-c by semi-preparative HPLC (3.0 mL/min, 210 nm,
ACN-H2O, 8.0:2.0, V/V). And Compound 11 (5.6 mg) and compound 12 were isolated from IXD-4-d
by semi-preparative HPLC (3.0 mL/min, 210 nm, ACN-H2O, 8.0:2.0, V/V).
2.3.1. Cyclocarioside Z1 (1)
White amorphous powder; [α]25 D + 8.1 (c 0.06, MeOH); HPLC-UV (ACN-H2O) λmax: 203
nm; 1H NMR (500 MHz in acetone-d6) data see Table 1, and 13C NMR (125 MHz in acetone-d6) data
see Table 3. HRESIMS, m/z 637.3956 [M – H]– (calcd for C35H57O10 637.3952).
2.3.2. Cyclocarioside Z2 (2)
White amorphous powder; [α]25 D – 10.6 (c 0.10, MeOH); HPLC-UV (ACN-H2O) λmax: 203
1
13
nm; H NMR (500 MHz in CD3OD) data see Table 1, and C NMR (125 MHz in CD3OD) data see
Table 3. HRESIMS, m/z 635.4157 [M – H]– (calcd for C36H59O9 635.4159).
2.3.3. Cyclocarioside Z3 (3)
White amorphous powder; [α]25 D – 87.5 (c 0.056, MeOH); HPLC-UV (ACN-H2O) λmax: 203
1
13
nm; H NMR (500 MHz in CD3OD) data see Table 1, and C NMR (125 MHz in CD3OD) data see
Table 3. HRESIMS, m/z 673.4293 [M + Na]+ (calcd for C37H62O9Na 673.4292).

2.3.5. Cyclocarioside Z4 (4)
White amorphous powder; [α]25 D + 32.0 (c 0.10, MeOH); HPLC-UV (ACN-H2O) λmax: 203
1
13
nm; H NMR (500 MHz in CD3OD) data see Table 1, and C NMR (125 MHz in CD3OD) data see
Table 3. HRESIMS, m/z 621.3997 [M – H]– (calcd for C35H57O9 622.4003).
2.3.6. Cyclocarioside Z5 (5)
White amorphous powder; [α]25 D – 87.6 (c 0.50, MeOH); HPLC-UV (ACN-H2O) λmax: 230
nm; 1H NMR (500 MHz in CD3OD) data see Table 2, and 13C NMR (125 MHz in CD3OD) data see
Table 3. HRESIMS, m/z 740.4947 [M + NH4]+ (calcd for C40H70NO11 740.4949).
2.3.7. Cyclocarioside Z6 (6)
Colorless powder; [α]25 D – 69.0 (c 0.10, MeOH); HPLC-UV (ACN-H2O) λmax: 230 nm; 1H
NMR (500 MHz in CD3OD) data see Table 2, and 13C NMR (125 MHz in CD3OD) data see Table 3.
HRESIMS, m/z 756.5256 [M + NH4]+ (calcd for C41H74NO11 756.5262).
2.3.8. Cyclocarioside Z7 (7)
Colorless powder; [α]25 D + 51.9 (c 0.027, MeOH); HPLC-UV (ACN-H2O) λmax: 230 nm; 1H
NMR (500 MHz in CD3OD) data see Table 2, and 13C NMR (125 MHz in CD3OD) data see Table 3.
HRESIMS, m/z 772.5215 [M + NH4]+ (calcd for C41H74NO12 772.5211).
2.3.9. Cyclocarioside Z8 (8)

1
Colorless powder; [α]25 D + 32.7 (c 0.07, MeOH); HPLC-UV (ACN-H2O) λmax: 230 nm; H
NMR (500 MHz in CD3OD) data see Table 2, and 13C NMR (125 MHz in CD3OD) data see Table 3.
HRESIMS, m/z 786.5369 [M + NH4]+ (calcd for C42H76NO12 786.5368).
2.3.10. Cyclocarioside I (9)
1
White amorphous powder; HPLC-UV (ACN-H2O) λmax: 230 nm; H NMR (500 MHz in
pyridine-d5) δH 2.93 (H-1a, 1H, m), 1.82 (H-1b, 1H, m), 1.46 (H-2a, 1H, m), 1.06 (H-2b, 1H, m), 2.09
(H-5, 1H, m), 1.29 (H-6, 2H, m), 1.43 (H-7a, 1H, m), 1.06 (H-7b, 1H, m), 2.09 (H-9, 1H, m), 2.88
(H-11a, 1H, m), 2.47 (H-11b, 1H, m), 4.00 (H-12, 1H, d, 7.5), 2.09 (H-13, 1H, m), 1.49 (H-15a, 1H,
m), 1.06 (H-15b, 1H, m), 1.99 (H-16, 2H, m), 1.98 (H-17, 1H, m), 0.88 (CH3-18, 3H, s), 1.15
(CH3-19, 3H, s), 1.47 (CH3-21, 3H, s), 2.65 (H-22a, 1H, m), 2.55 (H-22b, 1H, m), 6.13 (H-23, 1H, m),
6.42 (H-24, 1H, m), 5.04 (H-26a, 1H, br s), 4.96 (H-26b, 1H, br s), 1.89 (CH3-27, 3H, s), 4.90 (H-28a,
1H, br s), 4.89 (H-28b, 1H, br s), 1.80 (CH3-29, 3H, s), 0.98 (CH3-30, 3H, s), 4.78 (H-1', 1H, d, 7.0),
4.44 (H-2', 1H, m), 4.47 (H-3', 1H, m), 4.19 (H-4', 1H, m), 4.49 (H-5'a, 1H, br s), 3.55 (H-5'b, 1H, d,
13
15.5); C NMR (125 MHz in pyridine-d5) δC 38.9 (C-1), 29.5 (C-2), 148.3 (C-4), 51.4 (C-5), 25.3
(C-6), 34.6 (C-7), 40.5 (C-8), 40.4 (C-9), 39.8 (C-10), 32.3 (C-11), 74.2 (C-12), 43.9 (C-13), 50.7
(C-14), 31.1 (C-15), 25.2 (C-16), 50.1 (C-17), 16.4 (C-18), 20.3 (C-19), 73.9 (C-20), 27.2 (C-21),
45.0 (C-22), 128.1 (C-23), 135.4 (C-24), 142.4 (C-25), 114.6 (C-26), 18.6 (C-27), 113.4 (C-28), 24.0
(C-29), 16.3 (C-30), 99.6 (C-1'), 72.1 (C-2'), 73.8 (C-3'), 69.5 (C-4'), 67.5 (C-5').
2.3.11. Cyclocarioside J (10)

1
White amorphous powder; HPLC-UV (ACN-H2O) λmax: 203 nm; H NMR (500 MHz in
acetone-d6) δH 2.50 (H-1a, 1H, m), 1.51 (H-1b, 1H, m), 2.62 (H-2a, 1H, m), 2.23 (H-2b, 1H, m), 2.02
(H-5, 1H, m), 1.32 (H-6, 2H, m), 1.60 (H-7a, 1H, m), 1.22 (H-7b, 1H, m), 1.87 (H-9, 1H, m), 2.52
(H-11a, 1H, m), 1.34 (H-11b, 1H, m), 4.17 (H-12, 1H, m), 1.89 (H-13, 1H, m), 1.40 (H-15a, 1H, m),
1.07 (H-15b, 1H, m), 1.68 (H-16a, 1H, m), 1.32 (H-16b, 1H, m), 1.85 (H-17, 1H, m), 1.08 (CH3-18,
3H, s), 1.11 (CH3-19, 3H, s), 1.17 (CH3-21, 3H, s), 1.57 (H-22a, 1H, m), 1.44 (H-22b, 1H, m), 1.68
(H-23a, 1H, m), 1.57 (H-23b, 1H, m), 3.91 (H-24, 1H, m), 4.95 (H-26a, 1H, br s), 4.78 (H-26b, 1H, br
s), 1.82 (CH3-27, 3H, s), 4.87 (H-28a, 1H, br s), 4.78 (H-28b, 1H, br s), 1.73 (CH3-29, 3H, s), 1.02
(CH3-30, 3H, s), 4.35 (H-1', 1H, d, 6.5), 3.54 (H-2', 1H, m), 3.54 (H-3', 1H, m), 3.89 (H-4', 1H, m),
13
4.03 (H-5'a, 1H, br s), 3.54 (H-5'b, 1H, m); C NMR (125 MHz in acetone-d6) δC 37.9 (C-1), 29.2
(C-2), 148.1 (C-4), 51.5 (C-5), 24.8 (C-6), 34.6 (C-7), 40.5 (C-8), 40.2 (C-9), 39.6 (C-10), 32.1
(C-11), 74.0 (C-12), 43.9 (C-13), 50.4 (C-14), 30.8 (C-15), 25.0 (C-16), 59.5 (C-17), 16.0 (C-18),
19.8 (C-19), 73.5 (C-20), 25.3 (C-21), 36.1 (C-22), 30.1 (C-23), 74.9 (C-24), 148.5 (C-25), 109.3
(C-26), 17.5 (C-27), 113.0 (C-28), 23.4 (C-29), 16.3 (C-30), 98.9 (C-1'), 71.2 (C-2'), 72.8 (C-3'), 68.7
(C-4'), 66.5 (C-5').
2.3.12. Pterocaryoside B (11)
1
White amorphous powder; HPLC-UV (ACN-H2O) λmax: 203 nm; H NMR (500 MHz in
acetone-d6) δH 2.53 (H-1a, 1H, m), 1.57 (H-1b, 1H, m), 2.65 (H-2a, 1H, m), 2.27 (H-2b, 1H, m), 1.83
(H-5, 1H, m), 1.33 (H-6, 2H, m), 1.42 (H-7a, 1H, m), 1.07 (H-7b, 1H, m), 1.88 (H-9, 1H, m), 2.47
(H-11a, 1H, m), 1.33 (H-11b, 1H, m), 4.19 (H-12, 1H, td, 4.5, 5.0, 4.5), 1.96 (H-13, 1H, m), 1.61
(H-15a, 1H, m), 1.20 (H-15b, 1H, dt, 3.2, 13.1), 1.88 (H-16a, 1H, m), 1.70 (H-16b, 1H, m), 2.10 (H-17,

1H, m), 1.11 (CH3-18, 3H, s), 0.99 (CH3-19, 3H, s), 1.17 (CH3-21, 3H, s), 2.20 (H-22, 2H, m), 5.72
(H-23, 1H, m), 5.68 (H-24, 1H, d, 15.6), 1.26 (CH3-26, 1H, s), 1.25 (CH3-27, 3H, s), 4.86 (H-28a, 1H,
br s), 4.75 (H-28b, 1H, d, 1.7), 1.80 (CH3-29, 3H, s), 1.08 (CH3-30, 3H, s), 4.33 (H-1', 1H, d, 7.0),
3.52 (H-2', 1H, m), 3.52 (H-3', 1H, m), 3.77 (H-4', 1H, m), 3.89 (H-5'a, 1H, dd, 1.8, 10.7), 3.54 (H-5'b,
1H, m); 13C NMR (125 MHz in acetone-d6) δC 38.2 (C-1), 30.2 (C-2), 177.6 (C-3), 148.9 (C-4), 49.8
(C-5), 25.4 (C-6), 31.5 (C-7), 40.3 (C-8), 40.8 (C-9), 41.3 (C-10), 33.1 (C-11), 74.9 (C-12), 44.7
(C-13), 51.2 (C-14), 35.3 (C-15), 25.7 (C-16), 52.3 (C-17), 20.4 (C-18), 16.7 (C-19), 74.5 (C-20),
26.9 (C-21), 44.8 (C-22), 123.0 (C-23), 142.4 (C-24), 70.1 (C-25), 30.1 (C-26), 30.2 (C-27), 113.8
(C-28), 24.0 (C-29), 16.8 (C-30), 100.0 (C-1'), 72.4 (C-2'), 73.8 (C-3'), 69.4 (C-4'), 67.1 (C-5').
2.3.13. Cyclocarioside F (12)
1
White amorphous powder; HPLC-UV (ACN-H2O) λmax: 203 nm; H NMR (500 MHz in
acetone-d6) δH 2.53 (H-1a, 1H, m), 1.57 (H-1b, 1H, m), 2.66 (H-2a, 1H, m), 2.28 (H-2b, 1H, m), 1.83
(H-5, 1H, m), 1.33 (H-6, 2H, m), 1.43 (H-7a, 1H, m), 1.07 (H-7b, 1H, m), 1.88 (H-9, 1H, m), 2.49
(H-11a, 1H, m), 1.74 (H-11b, 1H, m), 4.19 (H-12, 1H, td, 4.6, 9.6), 1.98 (H-13, 1H, d, 11.1), 1.61
(H-15a, 1H, m), 1.24 (H-15b, 1H, m), 1.88 (H-16a, 1H, m), 1.74 (H-16b, 1H, m), 2.10 (H-17, 1H, m),
1.11 (CH3-18, 3H, s), 0.99 (CH3-19, 3H, s), 1.17 (CH3-21, 3H, s), 2.25 (H-22, 2H, d, 7.1), 5.71 (H-23,
1H, m), 5.46 (H-24, 1H, d, 15.8), 1.26 (CH3-26, 3H, s), 1.25 (CH3-27, 3H, s), 4.87 (H-28a, 1H, br s),
4.76 (H-28b, 1H, br s), 1.80 (CH3-29, 3H, s), 1.08 (CH3-30, 3H, s), 4.33 (H-1', 1H, d, 6.6), 3.52 (H-2',
1H, m), 3.52 (H-3', 1H, m), 3.77 (H-4', 1H, m), 3.89 (H-5'a, 1H, d, 12.6), 3.54 (H-5'b, 1H, m), 3.11
(OCH3, 3H, s); 13C NMR (125 MHz in acetone-d6) δC 38.4 (C-1), 30.4 (C-2), 149.0 (C-4), 52.3 (C-5),
25.8 (C-6), 35.4 (C-7), 41.4 (C-8), 40.8 (C-9), 40.4 (C-10), 33.0 (C-11), 74.8 (C-12), 44.8 (C-13),

51.3 (C-14), 31.6 (C-15), 26.5 (C-16), 50.1 (C-17), 16.9 (C-18), 20.5 (C-19), 74.4 (C-20), 27.1
(C-21), 45.0 (C-22), 127.3 (C-23), 138.9 (C-24), 75.1 (C-25), 26.5 (C-26), 26.1 (C-27), 113.8 (C-28),
24.0 (C-29), 16.8 (C-30), 100.0 (C-1'), 72.4 (C-2'), 73.8 (C-3'), 69.5 (C-4'), 67.2 (C-5').
2.4. 13C NMR calculation
Confab was used to search the conformational space of compound 1a and 1b. The obtained
stable conformers were optimized on B3LYP-D3(BJ)/SVP (IEFPCM, acetone) level of theory,
followed with frequency calculation. The conformers with Boltzmann population more than 1% were
further subjected to GIAO 13C NMR calculation on mPW1PW91/6-31G* (IEFPCM, acetone) level of
theory. The calculated shielding tensors of 1a and 1b were linearly regressed with experimental data,
respectively. The resulted linear formulas were used to convert shielding tensors into calculated
chemical shifts.
2.5. Acid hydrolysis
The configurations of sugar moieties were established according to the published method with
some modifications [14, 15]. Compounds 1-8 (each 0.5 mg) were hydrolyzed in 2 M HCl under reflux
in the oil bath at 100 ℃ for 3 h. The reaction mixture was neutralized by Na2CO3 and extracted with
CHCl3. The aqueous layer was concentrated and dried to obtain the monosaccharide fraction. The
residue was dissolved in pyridine (0.5 mL) containing 1 mg of L-cysteine methyl ester hydrochloride
and heated in the water bath at 60 ℃ for 1 h. Then trimethylsilylimidazole (0.5 mL) was added and
then heated at 60 ℃ for another 30 minutes. The reaction mixture was analyzed by GC-MS under the
conditions: Column, Rxi-5Sil MS (0.25 µm × 30.0 mm, 0.32 mm); front inlet 300 ℃, column 150

℃-300 ℃ at 15 ℃/min. The sugar configurations of compounds 1-8 were identified by the
comparison of the retention times with authentic standard treated by the same means.
2.6. Cell culture and treatment
The 3T3-L1 preadipocytes were gained from ATCC (Manassas, VA, USA). 3T3-L1
preadipocytes were grown in the complete medium containing DMEM supplemented with 10% FBS
and 1% penicillin-streptomycin at 37 ℃ under a humidified 5% CO2 incubator. The cells were
subcultured every 3-4 days to reach confluency, and then were seeded into 48-well plates at a density
of 2 × 104 cells/well. After 90% confluency, the cells were incubated with DMEM supplied with 10%
FBS and DMI (1 µM dexamethasone, 0.5 mM 3-isobutyl-1-methylxanthine, and 10 µg/mL insulin)
for 2 days. On the third day, differentiating media was replaced with a complete medium containing
10 µg/mL insulin and incubated for another 2 days (the fourth day). Thereafter, the cells were
maintained in complete medium for additional 6 days (the tenth day) with medium changing every 2
days. On day 10, fully differentiated adipocytes were treated with compounds at indicated
concentrations for 48 h.
2.7. Cell viability assay
Cell viability was determined by the MTT assay [16]. Briefly, 3T3-L1 adipocytes were seeded in
96-well plates at a density of 104 cells/well. And the cells were then treated with compounds at
indicated concentrations (3.125 µm-100 µm) for 48 h, respectively. Subsequently, each well was
added 20 µl MTT solution (5 mg/ml) and continued to culture at 37 ℃ for 4 h, followed by dissolving
the violet-formazan crystals with 150 µL DMSO. The absorbance at 490 nm was then measured with

a microplate reader. Cell viability of each group was expressed as a percentage of the control group
treated with DMSO.
2.8. Glucose consumption assay
Glucose consumption assay of 3T3-L1 adipocytes were measured by GOD-POD Kit as
described previously [17]. Fully differentiated adipocytes were treated with compounds at indicated
concentrations for 48 h. Then the glucose consumption of each well was obtained by subtracting the
glucose concentration of the cells from the glucose in the blank wells.
2.9. Statistical analysis
All results and data were confirmed at least three separate experiments. SPSS (Statistical
Package for the Social Sciences) software was used to determine significant differences via ANOVA
followed by Student's t-test. The data were expressed as means ± SD, and values P < 0.05 were
considered as statistically significant.
Table 1
1H NMR data of 1-4 (δ in ppm, J in Hz).
Position
1
1a
2.54, m
2b
3b
4b
2.44, td (13.5, 4.5)
1.60, m
2.48, td (13.5, 5.5)
1.46, m
1.63, m
1.58, m
2
5
2.66, m
2.65, m
2.75, td (13.0, 5.0)
2.29, m
2.36, m
2.22, m
2.28, td (12.0, 10.5)
2.09, overlapped
2.23, t (12.5, 11.5)
2.07, dd (13.0, 2.5)
2.03, d (12.5)
2.08, dd (13, 3.0)

6
7
1.78, m
1.34, m
1.61, m
1.34, m
1.90, m
1.36, m
1.60, m
1.22, m
1.94, m
2.39, m
1.41, m
4.08,
1.90, m
1.39, m
1.63, m
1.60, m
1.18, dd (11.5, 9.0)
1.21, overlapped
1.99, d (11.0)
2.50, m
1.23, m
9
1.95, overlapped
2.37, m
1.95, m
1.31, m
11
1.37, m
1.42, m
12
4.17,
4.12,
3.51,
td (10.5, 4.5)
1.74, m
td (10.5, 4.0)
1.91, m
td (10.5, 4.0)
1.80, overlapped
1.45, m
td (10.5, 5.0)
1.83, m
1.46, m
1.09, overlapped
1.82, m
1.64, m
1.80, m
1.10, s
13
15
1.43, m
1.60, m
1.09, overlapped
1.89, m
1.10, overlapped
1.82, m
1.30, m
16
1.90, m
1.64, m
1.62, m
1.56, m
17
18
19
21
22
1.88, m
1.82, m
2.55, m
1.09, s
1.08, s
0.97, s
1.12, s
1.11, s
1.08, s
0.90, s
1.31, s
1.17, s
1.19, s
1.22, s
1.89, m
1.62, m
2.23, m
1.70, m
1.51, m
1.43, m
1.40, m
23
3.13, m
1.67, m
5.68, m
1.86, m
1.60, m
1.72, overlapped
3.99, t (5.5, 7.0)
4.91, overlapped
4.81, br s
1.72, s
24
26
2.71, d (2.0)
1.22, s
4.00, t (6.0, 5.5)
4.96, overlapped
4.85, d (7.5)
1.75, s
5.67, m
1.29, br s
27
1.20, s
1.29, br s

28
4.87, br s
4.77, br s
1.80, s
4.86, d (7.5)
4.75, br s
1.80, s
4.84, overlapped
4.70, br s
1.77, s
4.87, overlapped
4.70, br s
29
30
1'
1.76, s
1.01, s
1.01, s
1.01, s
1.09, s
4.36, d (6.0)
3.53, m
4.38, d (8.0)
3.23, m
4.36, d (7.5)
3.21, m
4.97, d (7.0)
3.71, m
2'
3'
3.53, m
3.31, m
3.29, m
3.65, d (6.5)
3.68, m
4'
3.80, br s
3.89, br d (12.5)
3.52, m
3.02, t (9.0, 9.0)
3.27, m
3.02, t (9.5, 9.0)
3.28, m
5'
3.79, dd (11.5, 3.0)
3.59, q (5.5)
6'
1.28, d (6.0)
1.28, d (6.1)
3.64, s
-OCH3
aMeasured in acetone-d6 at 500 MHz.
bMeasured in CD3OD at 500 MHz.

Table 2
1H NMR (500 MHZ) data of 5-8 in CD3OD (δ in ppm, J in Hz).
Position
1
5
2.50, m
6
7
8
2.43, m
2.44, m
1.48, m
1.48, m
2.44, m
1.51, m
1.48, m
1.48, m
1.51, m
2
3
1.78, m
1.70, m
1.62, m
1.58, m
3.34, overlapped
3.34, overlapped
3.33, overlapped
3.34, overlapped

5
6
1.32, m
1.86, overlapped
1.45, m
1.35, m
1.86, m
1.46, m
1.60, m
1.23, m
1.80, m
2.34, m
1.40, m
4.11, td (10.5, 5.0)
1.79, m
1.45, m
1.07, m
1.79, m
1.61, m
1.80, m
1.00, s
1.35, m
1.46, m
1.35, m
1.45, m
7
1.59, m
1.60, m
1.23, m
1.61, m
1.23, m
1.23, m
9
1.80, m
1.82, m
1.81, m
11
2.39, m
2.42, m
2.44, m
1.38, m
1.36, m
1.34, m
12
13
15
4.12, td (10.5, 5.0)
1.82, m
4.07, td (13.5, 6.5)
1.70, m
4.07, td (10.5, 5.0)
1.70, m
1.45, m
1.47, m
1.46, m
1.07, m
1.09, overlapped
1.86, m
1.09, overlapped
1.99, m
16
1.79, m
1.61, m
1.82, m
17
18
19
21
22
1.80, m
1.90, m
0.99, s
1.11, s
1.17, s
2.41, m
1.70, m
1.47, m
3.81, m
1.17, s
1.22, s
0.94, s
0.89, s
1.01, s
1.91, m
1.02, s
1.02, s
1.12, s
1.12, s
1.11, s
1.18, s
1.18, s
1.17, s
2.32, m
1.48, m
2.41, m
1.70, m
23
24
26
27
28
29
30
5.77, m
6.20, d (15.5)
4.89, overlapped
1.86, s
2.07, m
5.15, t (7.5, 7.5)
1.70, s
1.46, m
3.80, t (7.5, 7.0)
1.17, s
1.65, s
1.22, s
0.98, s
0.98, s
0.94, s
0.90, s
0.90, s
0.89, s
0.99, s
1.00, s
1.01, s

1'
2'
3'
4'
5'
4.29, d (6.0)
3.61, m
4.29, d (6.0)
3.60, m
4.23, d (9.0)
3.21, m
4.25, d (8.0)
3.22, m
3.48, m
3.58, m
3.29, m
3.30, d (9.5)
2.99, t(9.5, 9.0)
3.26, m
3.83, m
3.84, m
3.27, m
3.89, m
3.86, m
3.87, m
3.53, m
3.52, m
3.16, m
6'
1.27, d (6.0)
4.35, d (8.0)
3.10, t (9.5, 7.5)
3.30, d (9.5)
3.02, t (10.0, 9.0)
3.26, m
1''
2''
3''
4''
5''
4.27, d (7.0)
3.46, m
4.33, d (7.5)
3.11, m
4.35, d (9.5)
3.11, m
3.58, m
3.29, m
3.29, m
3.79, m
2.99, t (9.0, 9.0)
3.27, m
2.99, t (11.5, 11.5)
3.27, m
3.86, m
3.51, m
6''
1.28, d (6.5)
1.27, d (7.5)
1.27, d (6.0)

Table 3
13C NMR data of 1-8 (δ in ppm).
Position
1
1a
2b
3b
4b
5b
6b
7b
8b
38.4
38.7
38.0
36.0
36.0
36.0
36.0
36.0

2
30.4
177.8
149.0
52.4
25.8
35.5
41.5
44.9
40.5
33.2
75.0
40.9
51.3
31.7
25.8
50.8
17.0
20.5
74.2
27.6
43.3
53.2
66.4
68.4
27.2
26.0
113.9
32.0
30.6
178.0
149.4
53.2
25.9
35.7
41.9
45.4
41.4
34.3
76.6
40.8
51.7
31.9
26.1
50.4
16.8
20.5
75.9
26.4
45.0
123.8
142.1
71.2
29.9
30.0
114.0
29.3
178.0
148.7
51.8
25.9
33.8
40.3
50.0
40.6
30.2
79.5
42.0
52.3
34.6
25.8
49.6
16.6
20.9
80.6
26.9
33.4
29.9
77.5
149.0
111.5
17.7
114.2
26.1
82.5
38.6
51.6
19.1
37.2
42.4
54.6
40.6
34.6
77.2
41.6
51.5
31.9
26.0
51.0
17.5
17.1
76.2
26.7
45.2
127.7
136.7
143.5
115.1
19.0
30.0
27.0
82.4
38.6
51.4
19.1
37.1
42.4
54.6
40.6
34.6
77.0
41.4
51.5
32.0
26.0
50.6
17.1
17.1
75.8
25.9
41.8
23.7
126.0
130.6
25.9
17.8
30.0
27.0
82.4
38.5
51.6
19.0
37.1
42.4
54.5
40.6
34.7
76.8
41.8
51.4
32.3
27.0
50.2
17.3
17.1
88.0
24.6
34.6
27.0
84.9
73.0
25.1
26.8
29.8
22.2
82.4
38.6
51.2
19.0
37.1
42.4
54.5
41.6
34.6
76.9
41.8
51.4
32.3
27.0
50.1
17.1
17.1
88.0
25.1
34.6
27.5
84.9
72.9
25.1
26.8
29.8
3
4
149.4
53.0
26.0
35.8
41.5
45.3
40.8
33.8
76.0
41.8
51.7
31.6
26.2
50.9
16.9
20.6
75.7
25.6
37.6
30.2
77.2
149.0
111.3
18.1
114.0
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28

29
24.0
17.0
100.1
72.4
73.9
69.5
67.3
24.0
16.9
99.8
75.3
77.7
77.1
72.6
18.0
24.0
17.0
100.6
75.4
77.8
77.2
73.0
18.1
23.7
15.7
96.5
71.4
73.1
72.4
65.0
23.3
17.2
101.5
72.5
74.7
69.3
67.6
23.2
17.4
101.5
72.5
74.3
69.3
66.1
23.2
17.3
102.2
75.0
78.1
71.5
66.9
23.2
17.3
101.4
75.4
78.1
77.3
73.0
18.1
100.7
75.6
78.0
77.2
73.0
18.1
30
1'
2'
3'
4'
5'
6'
1''
102.0
72.8
74.4
70.3
66.1
100.8
75.6
78.0
77.3
73.0
18.1
100.8
75.7
78.0
77.2
72.9
18.1
2''
3''
4''
5''
6''
-OCH3
52.0
aMeasured in acetone-d6 at 125 MHz.
bMeasured in CD3OD at 125 MHz.

3. Result and discussion
Compound 1 was isolated as a white amorphous powder, which had a molecular formula of
C35H58O10 determined by the HRESIMS ion peak at 637.3956 [M – H]– (calcd for C35H57O10
1
637.3952). The H NMR of compound 1 (Table 1) showed the presence of seven methyl proton
signals at δH 1.01 (s, H-30), δH 1.80 (s, H-29), δH 1.20 (s, H-27), δH 1.22 (s, H-26), δH 1.30 (s, H-21),
δH 1.11 (s, H-19), and δH 1.09 (s, H-18). Moreover, there were multiplets of methylene and methine
signals ranging from δH 1.00 to 2.80, as well as the signals of one terminal methylene olefinic protons
at δH 4.87 (br s, H-28a) and δH 4.77 (br s, H-28b), and an oxymethine proton at δH 4.17 (td, 1H, J =
10.5, 4.0 Hz, H-12). These data were used to assign the triterpenoid aglycone skeleton. The 13C NMR
(Table 3) showed 35 carbon signals, including signals of one terminal olefinic carbon at δC 149.0 and
δC 113.9, as well as five glycosyl carbon signals at δC 100.1, 72.4, 73.9, 69.5, and 67.3 ppm.
Therefore, an unusual 3,4-seco-dammarane type triterpenoid glycoside structure can be established
for compound 1, which is similar to known compound pterocaryoside B [19] except for the signals of
side-chain. In the HMBC spectrum, key correlations from H2-22 to C-23, H2-24 to C-23 and C-25,
from H3-21 to C-17, C-20, and C-22, from H3-26 to C-24 and C-25, together with chemical shifts of
C-23 and C-24 moved to low field verified the 23, 24-epoxy linkage and an additional hydroxyl group
linked to C-25. Furthermore, a pivotal HMBC correlation between the anomeric hydrogen H-1' and
C-12 determined the position of glycosyl. Besides, the glycosyl moiety was identified as
ɑ-L-arabinopyranosyl based on the NMR data, as well as further acid hydrolysis and GC-MS analysis
with L-arabinose standard (Table 5). The relative configurations of 1 were determined by NOESY
spectrum (Fig. 2). Strong correlations of H-5/H-9, H-9/H3-30, and H-12/H-17/H3-30 indicated that

H-5, H-9, H-12, H-17, and H3-30 are ɑ-oriented. The β-orientation of H-13, H3-18, and H3-19 were
then confirmed by NOESY correlations of H-13/H3-18 and H3-18/H3-19. The configuration at C-20
was determined to be S based on NOESY correlations of H-17/H3-21, respectively, on the basis of
comparison to the 13C NMR chemical shift for analogoues of 3,4-seco-dammaranes [18-20]. The key
NOESY correlations between H3-21 and H-23, H3-26 and H-22a, H3-26 and H-23 indicated that H-23
and H-24 were trans oriented. Therefore, there were two possible configurations, namely 23S24R (1a)
and 23R24S (1b). To determine the absolute configuration of the epoxy group, we performed GIAO
13
DFT C NMR calculations of structure 1a and 1b, and the mean absolute deviation (MAD) and
sDP4+ values [21] of them showed that 1a was the correct structure (Table 4). From all the evidence
above, the chemical structure of
1
was established as (12R, 20S, 23S,
24R)-20,25-dihydroxy-23,24-epoxy-12-O-α-L-arabinopyranosyl-3,4-secodammara-4(28)-en-3-oic
acid.

Table 4
GIAO DFT calculation of 13C NMR data of structures of 1a and 1b (δ in ppm).
Position
Exptl.
38.4
30.4
177.8
149.0
52.4
25.8
35.5
41.5
44.9
40.5
33.2
75.0
40.9
51.3
31.7
25.8
50.8
17.0
20.5
Calcd. of 1a
37.3
Deviation a
1.1
Calcd. of 1b
36.5
Deviation b
1.9
1
2
31.0
0.6
29.4
1.0
3
176.4
149.4
49.8
1.4
174.5
148.2
49.4
3.3
4
0.4
0.8
5
2.7
3.1
6
25.5
0.5
25.3
0.7
7
35.1
0.4
34.0
1.5
8
41.4
0.1
40.5
1.0
9
40.3
0.6
40.5
0.4
10
11
12
13
14
15
16
17
18
19
42.5
2.0
41.4
0.9
27.2
6.0
28.0
5.2
73.0
2.0
73.2
1.8
48.5
3.6
48.6
3.7
53.7
2.4
54.4
3.1
32.4
0.6
34.3
2.5
26.1
0.3
28.7
2.9
52.0
1.2
45.9
4.9
17.3
0.4
16.2
0.7
21.9
1.4
21.8
1.3

20
21
22
23
24
25
26
27
28
29
30
1'
74.2
27.6
43.3
53.2
66.4
68.4
27.2
26.0
113.9
24.0
17.0
100.1
72.4
73.9
69.5
67.3
74.2
25.3
43.8
54.2
62.9
71.5
26.7
26.7
116.7
25.1
18.0
97.1
71.9
73.4
70.8
68.1
MAD
sDP4+
0.0
2.3
0.5
0.9
3.6
3.1
0.5
0.7
2.8
1.0
1.1
3.0
0.5
0.5
1.3
0.8
1.5
74.6
26.3
44.6
55.8
68.0
67.3
28.0
28.0
115.7
24.7
19.2
96.3
70.9
72.9
69.0
67.5
MAD
sDP4+
0.4
1.3
1.3
2.5
1.5
1.1
0.8
2.0
1.8
0.6
2.3
3.8
1.5
1.0
0.5
0.2
1.8
0%
2'
3'
4'
5'
100%
Compound 2 was obtained as a white amorphous powder. The molecular formula of 2 was
determined to be C36H60O9 by HRESIMS. The 1D, 2D NMR data were almost identical with
cyclocarioside J [18] except for the replacement of the arabinopyranosyl moiety by a
quinovopyranosyl unit in 2. The HMBC (Fig. 2) correlations between δH 4.38 (d, J = 8.0 Hz, H-1')
and δC 76.0 (C-12) indicated that the glycosyl was located at C-12. Moreover, further acid hydrolysis
and GC-MS analysis with a D-quinovose standard (Table 5) suggested a quinovopyranosyl unit in 2.
Therefore, the structure of 2 was deduced as (12R, 20S, 24S)-20,24-dihydroxy-12-O-β-D-

quinovopyranosyl-3,4-secodammara-4(28),25-dien-3-oic acid.
Compound 3 was obtained as a white amorphous powder. The molecular formula of 3 was
determined to be C37H62O9 by HRESIMS. The 1D and 2D NMR data were almost identical to known
compound pterocaryoside A [19] except for an additional methoxy group, as indicated a methyl
proton signal at δH 3.64 (s, OCH3) and a carbon signal at δC 52.0 (OCH3). Moreover, the key
correlation between OCH3 and C-3 in the HMBC (Fig. 2) revealed that 3 was a 3-methyl ester
derivative of pterocaryoside A. Besides, the glycosyl moiety was determined by acid hydrolysis and
GC-MS analysis with D-quinovose standard (Table 5). Therefore, the structure of 3 was deduced as
(23E)-(12R,
20S)-12,20,25-trihydroxy-12-O-β-D-quinovopyranosyl-3,4-secodammara-4(28),23-dien-3-oic
acid
methyl ester.
Compound 4 has the molecular formula C35H58O9, as determined by HRESIMS analysis (m/z
621.3997 [M – H]–). Detailed analysis of NMR spectra indicated that 4 was a 3,4-seco-dammarane
triterpenoid glycoside similar to cyclocaroside O [22]. However, the 5-O-acetyl-ɑ-L-arabinofuranosy
at C-20 in cyclocaroside O was replaced by a ribopyranosyl moiety in 5. And the linkage of glycosyl
fragment was confirmed by a signficant HMBC correlation between δH 4.99 (d, J = 7.0 Hz, H-1') and
δC 80.5 (C-20). Besides, the ribose was identified by acid hydrolysis and GC-MS analysis with
standard
(Table
5).
Thus
compound
4
was
characterized
as
(12R,
20S,
24S)-12,20,24-trihydroxy-20-O-β-D-ribopyranosyl-3,4-secodammara-4(28),25-dien-3-oic acid.
Compound 5 was obtained as a white amorphous powder. The molecular formula of 5 was

determined to be C40H66O11 by HRESIMS. Full NMR data anlysis indicated that 5 was a triterpenoid
glycoside which similar to cyclocarioside K [18]. In the 1H NMR (Table 2), there were seven methyl
proton signals (δH 1.86, 1.18, 1.12, 1.02, 0.99, 0.98, 0.90) assigned to the aglycone moiety, together
with two anomeric proton signals at δH 4.29 (d, 1H, J = 6.0 Hz) and δH 4.27 (d, 1H, J = 10.7 Hz) for
13
sugar moieties in the molecule. Detailed analysis of the C NMR (Table 3) data revealed that there
were two anomeric carbon signals at δC 101.5 (C-1') and δC 102.0 (C-1''), which also proved the
presence of two sugar moieties in 5. The structure of two sugar moieties was determined to be
arabinose by acid hydrolysis and GC-MS analysis with standard (Table 5). The correlations from
H-1' to C-3, H-1'' to C-12 in the HMBC spectrum (Fig. 2) confirmed the linkage of glycosyl fragment
located at C-3 and C-12. In the NOESY spectrum (Fig. 2), correlations between H-3 (δH 3.34), H-1β
(δH 1.48, β-orientation) and H-29 (δH 0.90), as well as correlations between H-12 (δH 4.12), H-11ɑ (δH
2.39, ɑ-orientation), and H-9 (δH 1.80) indicated the H-3, H-12 were a-orientation and β-orientation.
Furthermore, the side-chain contained one terminal methylene olefinic proton signals at δH 4.89 (m,
H-26), together with a pair of olefinic proton signals at δH 5.77 (m, H-23) and 6.20 (d, 1H, J = 15.6
Hz, H-24), whose the coupling constant (J = 15.6 Hz) suggested that the configuration was E. The
13
absolute configuration at C-20 was determined to be S based on the C NMR spectra by comparing
chemical shifts with analogous dammaranes [18, 23]. Consequently, the compound 5 was determined
as
(23E)-(20S)-(3α,
12β)-20-hydroxy-dammara-23,25-dien-12-O-α-L-arabinopyranoside-3-O-α-L-arabinopyranoside.
Compound 6 was isolated as colorless powder, which had a molecular formula of C41H70O11
determined by the HRESIMS. Detailed analysis of 1D and 2D NMR data showed that 6 had a high

similarity to cyclocarioside K except for the only distinction in side-chain. Therefor, the structure of
side-chain was futher confirmed by the correlations from H3-21 to C-17, C-20, C-22 and C-23, H3-26
to C-24, and C-25, H3-27 to C-24, and C-25, and H2-23 to C-24 in the HMBC spectrum. So the
structure
of
6
was
deduced
as
(20S)-(3α,
12β)-20-hydroxy-dammara-24-dien-12-O-β-D-quinovopyranoside-3-O-α-L-arabinopyranoside.
Compound 7 was isolated as colorless powder, which had a molecular formula of C41H70O12
determined by the HRESIMS. Detailed analysis of NMR data showed that 7 had a high similarity to
cyclocarioside K except for replacement of arabinopyranosyl moiety at C-3 by a xylopyranosyl
moiety at the same position in 7. Moreover, xylopyranosyl moiety was determined by acid hydrolysis
and GC-MS analysis with standard (Table 5). Therefore, the structure of 7 was deduced as (20S,
24R)-(3α,
12β)-20,24-epoxydammara-25-hydroxyl-12-O-β-D-quinovopyranoside-3-O-β-D-xylopyranoside.
Compound 8 was obtained as colorless powder. The molecular formula of 8 was determined to
be C42H72O12 by HRESIMS. The 1D and 2D NMR data were almost identical to known compound
cyclocarioside K except for the only difference with the glycosyl moiety at C-3. There was a
D-quinovose located in C-3, which was confirmed by acid hydrolysis and GC-MS analysis with
standard (Table 5). Thus, the compound
8
was determined as (20S, 24R)-(3α,
12β)-20,24-epoxydammara-25-ol-12-O-β-D-quinovopyranoside-3-O-β-D-quinovopyranoside.
Cell viability was determined using MTT assay. The cell viability above 90% were considered as
non-cytotoxicity. As shown in Table 6, the maximum safe dosages for most compounds were 50µM