The phytochemical investigation of the hexane extract from the twigs and leaves of Dacrycarpus imbricatus (Blume) de Laub led to the isolation of a rare sesquiterpene, spathulenol (1) along with three diterpenes named pimaric acid (2), trans-communic acid (3) and cis-communic acid (4). Their structures were determined by combination of spectral analysis and comparison with reported data. This is the first report on isolation of compound 1 from the Podocarpaceae family. Keywords. Dacrycarpus imbricatus, spathulenol, pimaric acid, communic acid

Hau, Dang Viet

Hoang Anh, Nguyen Thi

Hong Nhun, Le Thi

Loc, Tran Van

Quan, Tran Duc

Sung, Tran Van

Tam, Nguyen Thanh

Thien, Dao Duc

Thuy, Trinh Thi

Tinh, Bui Xuan

Tuan, Nguyen Le

Vietnam Academy of Science and Technology: Journals Online

  
Vietnam Journal of Chemistry, International Edition, 55(6): 734-737, 2017 
DOI: 10.15625/2525-2321.2017-00536 
734 
 
 
 
Terpenoids from Dacrycarpus imbricatus 
Dang Viet Hau
1,2
, Nguyen Thanh Tam
1,2
, Nguyen Thi Hoang Anh
1,2
, Tran Duc Quan
1
,  
Dao Duc Thien
1
, Le Thi Hong Nhung
3
, Bui Xuan Tinh
4
, Nguyen Le Tuan
5
, Tran Van Loc
1
,  
Tran Van Sung
1
, Trinh Thi Thuy
1,2*
 
1Institute of Chemistry, Vietnam Academy of Science and Technology (VAST) 
2Graduate University of Science and Technology, VAST 
3University of Industry 
4Quang Trung University 
5Qui Nhon University 
Received 19 October 2017; Accepted for publication 29 December 2017 
 
Abstract 
The phytochemical investigation of the hexane extract from the twigs and leaves of Dacrycarpus imbricatus 
(Blume) de Laub led to the isolation of a rare sesquiterpene, spathulenol (1) along with three diterpenes named pimaric 
acid (2), trans-communic acid (3) and cis-communic acid (4). Their structures were determined by combination of 
spectral analysis and comparison with reported data. This is the first report on isolation of compound 1 from the 
Podocarpaceae family.  
Keywords. Dacrycarpus imbricatus, spathulenol, pimaric acid, communic acid.  
 
1. INTRODUCTION 
 
Dacrycarpus imbricatus (Blume) de Laub 
(Podocarpus kawaii Hayata) is a coniferous tree, up 
to 40 m tall with red brown bark, belonging to 
Podocarpaceae family. It is mainly distributed in the 
West Pacific islands near China and Viet Nam. This 
tree is classified as 'Least Concern' in the IUCN Red 
List of Threatened Species (2011) [1]. Its timber had 
been used in construction, for furniture, firewood 
and in some areas cultivated as ornamental tree. In 
our previous paper, we described the isolation and 
identification of 20-hydroxyecdysone from the bark 
of D. imbricatus and its effect on OCI-AML cell 
proliferation in vitro [2]. In the course of our 
continuing search for the bioactive constituents of 
this plant, a rare sesquiterpene, spathulenol (1) and 
three diterpenes:pimaric acid (2), mixture of trans- 
and cis-communic acid (3+4) were isolated from the 
twigs and leaves of D. imbricatus. Their structures 
were elucidated by the spectroscopic means and 
comparison with published data.  
 
2. EXPERIMENTAL 
 
2.1. Materials and methods 
 
D. imbricatus was collected in Lam Dong province, 
Viet Nam (August, 2012) and identified by Dr. 
Nguyen Tien Hiep (Vietnam National Museum of 
Nature). A voucher specimen (VNMN. 
B0000050010) was deposited in the Vietnam 
National Museum of Nature, Vietnam Academy of 
Science and Technology (VAST). 
ESI-MS: Agilent LC-MSD-Trap SL. 1H-NMR 
(500.13 MHz) and 13C-NMR (125.77 MHz) spectral 
data were measured on a Bruker Avance 500 
Ultrashield NMR Spectrometer at 25 oC. Chemical 
shifts were reported as  values with reference to 
TMS as internal standard for 1H ( = 0 ppm) and 
CDCl3 for 
13C ( = 77.0 ppm). TLC: Silica gel 60 
F254 (0.25mm, Merck); reversed phase RP18F254S 
(0.25mm, Merck). CC: Silica gel 60 (230-400 mesh, 
Merck) for the first column, silica gel 60, 40-63 m 
(Merck) for the following columns. 
 
2.2. Extraction and isolation 
 
The dried ground twigs and leaves of D. imbricatus 
(900 g) were extracted with methanol-water (90:10, 
v/v) at room temperature (4 times x 3 lit, each 
overnight). After concentration under reduced 
pressure, the obtained crude extract (62.2 g) was 
  
VJC, 55(6), 2017                                           Trinh Thi Thuy et al. 
735 
suspended in water (150 ml) and then sequentially 
partitioned with n-hexane, ethyl acetate and n-
butanol. The organic solvents were evaporated to 
yield the corresponding extracts (10.8 g, 18.0 g and 
10.2 g, respectively). The condensed n-hexane 
extract (10.8 g) was separated on silica gel column, 
eluting with a gradient of n-hexane-EtOAc (100:0  
80:20) to yield 40 fractions (FH1–FH40) on the 
basis of TLC analysis. Fraction FH 22 was further 
purified on a silica gel column eluted with n-hexane-
EtOAc (6:4, v/v) to furnish compound 1 (10 mg). 
Recrystallization of the precipitated solid from 
fraction FH 31 with cold methanol gave compound 2 
(30 mg). The mixture of compounds 3+4 (20 mg) 
was obtained from FH3 by column chromatography 
on silica gel eluted with hexane:EtOAc (9:1  1:1). 
 
2.3. Spectral data of isolated compounds 
 
Spathulenol (1): Colorless semisolid. ESI-MS m/z: 
221.3 [M+H]+. 1H-NMR (CDCl3, 500 MHz): 4.69 
(1H, s, H-14a), 4.66 (1H, s, H-14b), 2.42 (1H, dd, J 
= 13.0, 6.0 Hz, H-4a), 2.20 (1H, m, H-6), 2.06 (1H, 
d, J = 13.0 Hz, H-4b), 1.77 (1H, m, H-8a), 1.55 (1H, 
m, H-8b), 1.32 (1H, d, J = 10.5 Hz, H-10), 1.28 (3H, 
s, H-15), 1.06 (3H, s, H-12), 1.03 (3H, s, H-13), 0.71 
(1H, ddd, J = 11.0, 9.5, 6.0 Hz, H-2), 0.47 (1H, dd, J 
= 11.0, 9.5 Hz, H-1). 13C-NMR (CDCl3, 125 MHz): 
153.46 (C-5), 106.26 (C-14), 80.99 (C-9), 54.35 (C-
10), 53.41 (C-6), 41.75 (C-8), 38.87 (C-4), 29.92 (C-
1), 28.66 (C-12), 27.50 (C-2), 26.72 (C-7), 26.08 (C-
15), 24.79 (C-3), 20.27 (C-11), 16.34 (C-13). 
Pimaric acid (2): Colorless semisolid. ESI-MS 
m/z: 303.6 [M+H]+. 1H-NMR (CDCl3, 500 MHz): 
5.77 (1H, dd, J = 17.0, 10.5 Hz, H-15), 5.22 (1H, s, 
H-14), 4.90 (1H, dd, J = 17.0, 1.5 Hz, H-16a), 4.88 
(1H, dd, J = 10.5, 1.5 Hz, H-16b), 1.21 (3H, s, H-
19), 1.04 (3H, s, H-17), 0.84 (3H, s, H-20). 13C-
NMR (CDCl3, 125 MHz): 184.92 (C-18), 148.91 (C-
15), 136.63 (C-8), 129.13 (C-14), 110.16 (C-16), 
50.58 (C-9), 48.84 (C-5), 47.28 (C-40, 38.30 (C-1), 
37.74 (C-10), 37.40 (C-13), 37.05 (C-3), 35.48 (C-
7), 34.46 (C-12), 26.04 (C-17), 24.91 (C-6), 18.57 
(C-11), 18.16 (C-2), 16.78 (C-19), 15.22 (C-20). 
Trans- and cis-communic acid (3+4): 
Semisolid. ESI-MS m/z: 303.1 [M+H]
+
, ratio of 
trans- and cis-isomers  4:1). 
Trans-communic acid (3): 1H-NMR (CDCl3, 500 
MHz): 6.32 (1H, dd, J = 17.0, 10.5 Hz, H-14), 5.41 
(1H, t, J = 6.0 Hz, H-12), 5.04 (1H, d, J = 17.0 Hz, 
H-15a), 4.88 (1H, d, J = 10.5 Hz, H-15b), 4.84 (1H, 
s, H-17a), 4.47 (1H, s, H-17b), 1.75 (3H, s, H-16), 
1.25 (3H, s, H-18), 0.65 (3H, s, H-20). 13C-NMR 
(CDCl3, 125 MHz): 183.90 (C-19), 147.93 (C-8), 
141.61 (C-12), 133.90 (C-14), 133.45 (C-13), 109.91 
(C-15), 107.66 (C-17), 56.43 (C-5), 56.29 (C-9), 
44.21 (C-4), 40.37 (C-10), 39.27 (C-6), 38.49 (C-1), 
37.93 (C-3), 29.04 (C-18), 25.84 (C-7), 23.31 (C-
11), 19.93 (C-2), 12.83 (C-20), 11.84 (C-16). 
Cis-communic acid (4): 1H-NMR (CDCl3, 500 
MHz): 6.78 (dd, J = 17.5, 11.0 Hz, H-14), 5.31 (t, J 
= 6.0 Hz, H-12), 5.18 (d, J = 17.5 Hz, H-15a), 5.08 
(d, J = 11.0 Hz, H-15b), 4.84 (s, H-17a), 4.49 (s, H-
17b), 1.77 (s, H-16), 1.25 (s, H-20), 0.65 (s, H-20). 
13C-NMR (CDCl3, 125 MHz): 183.90 (C-19), 147.93 
(C-8), 133.86 (C-14), 131.67 (C-12), 131.56 (C-13), 
113.25 (C-15), 107.80 (C-17), 56.70 (C-9), 56.43 
(C-5), 44.21 (C-4), 40.43 (C-10), 39.27 (C-6), 38.52 
(C-1), 37.93 (C-3), 29.04 (C-18), 25.84 (C-7), 22.28 
(C-11), 19.93 (C-2), 19.72 (C-16), 12.83 (C-20). 
 
2.4. Bioactivity tests 
 
Cytotoxicity tests were performed according to 
Likhiwitayawuid et al. [9] and Skehan et al. [10] at 
different concentrations in 96-well plates. The KB 
and Hep-G2 cell lines were maintained in RPMI-
1640 culture medium with 10 % fetal bovine serum 
(FBS). Ellipticine was used as positive control. 
 
 
Figure 1: Chemical structures of isolated 
compounds 1–4 from Dacrycarpus imbricatus 
 
3. RESULTS AND DISCUSSION 
 
Compound 1 was obtained as a colorless semisolid. 
Its molecular formula, C15H24O, was deduced from 
combined analysis of ion peak at m/z 221.3 [M+H]+ 
in the positive ESI MS and 1H-, 13C-NMR and 
DEPT spectral data. 13C-NMR and HSQC spectral 
data of 1 confirmed the presence of 15 carbons, 
including three methyls, five methylenes, four 
methines and three quaternary carbons (in which has 
  
VJC, 55(6), 2017        Terpenoids from Dacrycarpus imbricatus. 
736 
one carbon bonded to oxygen atom). The 1H-NMR 
spectrum showed the presence of three methyl 
groups bonded to quaternary carbons at δH 1.28 (3H, 
s, H-15), 1.06 (3H, s, H-12), 1.03 (3H, s, H-13), an 
exocyclic methylene at δH 4.69 (1H, s, H-14a), 4.66 
(1H, s, H-14b), a cyclopropyl moiety at δH 0.71 (1H, 
ddd, J = 11.0, 9.5, 6.0 Hz, H-2), 0.47 (1H, dd, J = 
11.0, 9.5 Hz, H-1). Two typical cyclopropyl protons 
are characteristic of an aromadendrane-type and 
these data are in accordance with a sesquiterpenoid 
[3, 4]. In the HMBC experiment, the correlations 
between H2-14 and C-4, C-5, C-6; H-10; H2-8 with 
C-9 was presented, suggesting the exocyclic 
methylene and OH groups was located at C-5, C-9, 
respectively. The nuclear overhauser effect (NOEs) 
between the methine proton H-1 and the methyl 
protons H-15 implied that the stereochemistry of the 
methyl and hydroxyl groups at C-9 were α- and β-
oriented, respectively. Combined all the above 
spectroscopic data and comparison with literature 
data [4], the structure of 1 is identified as 
spathulenol. This compound is a rare sesquiterpene 
which was previously isolated from the plant 
Eucalyptus spathulata [5], but this is the first time it 
has been found in the Podocarpaceae family [3-5].  
Compound 2 was isolated as a semisolid. The 
molecular formula of 2 was assigned as C20H30O2 by 
the molecular ion peak at m/z 303.2 [M+H]+ in the 
positive ESI-MS and NMR data. 1H NMR spectrum 
of 2 showed a terminal vinyl group [δH 5.77 (1H, dd, 
J = 17.0, 10.5 Hz, H-15), 4.90 (1H, dd, J = 17.0, 1.5 
Hz, H-16a), 4.88 (1H, dd, J = 10.5, 1.5 Hz, H-16b); 
δC 148.91 (C-15), 110.16 (C-16)], a trisubstitued 
olefin group (δH 5.22 (1H, s, H-14); δC 136.63 (C-8), 
129.13 (C-14)] and three methyl groups at δH 1.47, 
1.60, 2.37 (each 3H, s). Except for these moieties the 
13C-NMR and HSQC showed the resonances of a 
carboxyl group at δC 184.94, seven methylenes, two 
methines and three quaternary carbons. These data 
suggested that 2 is a pimarane diterpenoid. By 
means of its spectrometric analysis and comparison 
with previously published data [6], compound 2 was 
determined as pimaric acid.  
Trans- and cis-communic acid (3+4) were 
isolated as a mixture. In the 1H- and 13C NMR 
spectra showed many quite similar pairs of signals in 
chemical shifts and shape of two compounds with a 
ratio of about 4:1. The ESI-MS showed one 
molecular ion peak at m/z 303.1, which indicated 
this is a mixture of two isomers. The 1H-NMR 
spectrum exhibited signals for three methyl groups 
at δH 1.75/1.77, 1.25 and 0.65/0.64 and an 
exomethylene at δH 4.84/4.87 and 4.47/4.49 (4×s), 
which were characteristic of a labdane-type 
diterpene. Additionally it exhibited downfield 
signals for an olefinic protons at δH 5.41/5.31 (t, J = 
6.0 Hz), a vinylic  group at δH 6.32 (1H, dd, J = 
17.0, 10.5 Hz/6.78 (dd, J = 17.5, 11.0 Hz); 5.04 (1H, 
d, J = 17.0 Hz), 4.88 (1H, d, J = 10.5 Hz)/5.18 (d, J 
= 17.5 Hz), 5.08 (d, J = 11.0 Hz). Comparison of 
their NMR data with those reported in the literatures 
[7, 8], 3 and 4 were determined as trans- and cis-
communic acid, respectively. 
The cytotoxic activity of of the major pure 
compound (2, 30 mg) was evaluated against the two 
human cancer cell lines KB (mouth epidermal 
carcinoma) and HepG2 (hepatocellular carcinoma). 
Compound 2 was medium active against both tested 
cancer cell lines with IC50 values of 173.8 and 99.6 
M, respectively.  
 
Acknowledgments. We would like to thank the 
Ministy of Science and Technology of Vietnam of 
Vietnam for financial support for this research (NDT 
2017-2019) and the Italian Ministero degli Esterie 
Cooperazione Internazionale (MAECI) for financial 
support. We thank Master Dang Vu Luong for the 
NMR spectra. 
 
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Corresponding author: Trinh Thi Thuy 
Institute of Chemistry 
Vietnam Academy of Science and Technology 
18, Hoang Quoc Viet road, Cau Giay district, Hanoi, Viet Nam 
E-mail: thuy@ich.vast.vn; Tel. 024.32121149/0978987562. 
 


Terpenoids from Dacrycarpus imbricatus.

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Terpenoids from Dacrycarpus imbricatus.

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