全 文 ：天然产物研究与开发 Nat Prod Ｒes Dev 2015，27:1007-1010
文章编号:1001-6880(2015)6-1007-04
Ｒeceived:December 16，2014 Accepted:April 28，2015
Foundation item:National Natural Science Foundation of China
(21262021)
* Corresponding author Tel:86-871-65920569;E-mail:rongtaolikm@
163． com
金刚纂的化学成分及其抗 HIV和抗肿瘤活性研究
李洪梅，王若菲，李蓉涛*
昆明理工大学 生命科学与技术学院，云南昆明 650500
摘 要:从金刚纂全草中分离得到一个新的天然来源的阿替生烷型二萜，3-methyl-agallochaol C(1)，以及 13 个
已知化合物。其中，化合物 4 和 9 具有一定的抗 HIV-1 活性，化合物 10 具有一定的抗肿瘤活性。
关键词:金刚纂;3-methyl-agallochaol C;抗 HIV;抗肿瘤
中图分类号:Ｒ284． 1 文献标识码:A DOI:10． 16333 / j． 1001-6880． 2015． 06． 012
Chemical Constituents from Euphorbia neriifolia and
Their Ｒelated Anti-HIV and Anti-cancer Activities
LI Hong-mei，WANG Ｒuo-fei，LI Ｒong-tao*
Faculty of Life Science and Technology，Kunming University of Science of Technology，Kunming 650500，China
Abstract:Systematic investigation on the whole plant of Euphorbia neriifolia led to the isolation of a new natural atisane
diterpenoid，3-methyl-agallochaol C (1)，along with thirteen known compounds (2-14)． Among them，compounds 4 and
9 exhibited moderate anti-HIV-1 activity，and compound 10 possessed moderate anti-cancer activity．
Key words:Euphorbia neriifolia;3-methyl-agallochaol C;anti-HIV;anti-cancer
Introduction
Human immunodeficiency virus (HIV)is the etiologic
agent of the acquired immunodeficiency syndrome
(AIDS)，a disease that already claimed the lives of
more than 25 million people． The global incidence of
HIV infection in 2010 was estimated to be approximate-
ly 33． 2 million people ［1］． Current antiretroviral drugs
are vitally important to improve the quality and prolong
the life of HIV /AIDS patients． Nevertheless，these
drugs have many disadvantages including resistance，
toxicity，limited availability，high cost and lack of any
curative effect ［2］． Thus，the need and demand has
prompted an intense research effort to discover new，se-
lective and safe drugs for the treatment of HIV /AIDS．
Natural sources，particularly plants，are an excellent
source of anti-HIV agents． Southern China，especially
Yunnan Province，possess an abundant plant biodiversi-
ty and a long history of medicinal use of plants，so man-
y plants may contain novel anti-HIV constituents．
Euphorbia neriifolia Linn．(Euphorbiaceae)，traditional
Dai medicine in China，is a landscape plant widely cul-
tivated in the south and southwest of Yunnan Province
and used for hedges ［3］． This plant produces milky la-
tex which possesses several applications in folk medi-
cines，such as irritant，emetic，purgative and diuretic
［4］． The plant extracts were demonstrated to exhibit an-
tihepatotoxic and cytotoxic activities ［5］． In order to dis-
cover anti-HIV agents of natural origin，different parts
(EtOAc，n-BuOH and H2O parts)of 95% EtOH ex-
tracts of E． neriifolia were evaluated for their anti-HIV-
1 activities，using AZT as positive control (EC50 =
0. 008 μg /mL)． Ｒesults showed that EtOAc part exhib-
ited potential anti-HIV-1 activity with an EC50 value of
1． 26 μg /mL． Bioassay-guided isolation of the EtOAc
part led to the purification of fourteen compounds (1-
14)，including a new natural atisane diterpenoid，3-
methyl-agallochaol C (1)． Among them，compounds 4
and 9 possessed moderate anti-HIV-1 activity． In addi-
tion，in view of the cytotoxic activity of this plant repor-
ted previously，the cytotoxicities of isolated compounds
were also tested．
Experimental
General
Optical rotation was run on a Jasco DIP-370 digital po-
larimeter (JASCO Corporation，Tokyo，Japan)． NMＲ
spectra were recorded over Bruker AM-400，DＲX-500
and AVANCE III-600 instruments with tetramethylsi-
lane (TMS)as an internal standard (Bruker BioSpin
Group，Germany)． ESI-MS was obtained with an API-
Qstar-TOF instrument． Column chromatography (CC)
was performed with silica gel (200-300 mesh，Qingdao
Marine Chemical and Industrial Factory，China)，MCI
(MCI-gel CHP-20P，75-150 μm，Mitsubishi Chemical
Corporation)and Sephadex LH-20 (Amersham Biosci-
ences AB，Uppsala，Sweden)． Fractions were monitored
by TLC plates (Si gel GF254，Qingdao Marine Chemical
and Industrial Factory，China)，and spots were visual-
ized by heating silica gel plates sprayed with 5%
H2SO4-EtOH．
Plant material
The whole plant of E． neriifolia was collected from
Xishuangbanna，Yunnan Province，PＲ China，in Sep-
tember 2008，and was identified by Mr． Jing-yun Cui． A
voucher specimen (No． 20080901)was deposited at
the Laboratory of Phytochemistry，Faculty of Life Sci-
ence and Technology，Kunming University of Science of
Technology．
Extraction and isolation
The air-dried and powdered woods of E． neriifolia (4． 0
kg)were extracted with 95% EtOH (3 × 6 L，24 h
each)at room temperature，and then concentrated un-
der vacuum to yield an extract (125 g)，which was
suspended in H2O (2 L)and then extracted with EtO-
Ac (4 × 2 L)． The EtOAc extract (66． 0 g)was sep-
arated by MCI，eluting with MeOH /H2O (gradient
30%，60%，90% and 100%)，to afford fractions A-E．
Fr． C (1． 6 g)was subjected to silica gel CC (200-300
mesh) ，using petroleum ether /acetone (5∶ 1)as eluent
to give six subfractions，C-1 ～ C-6． Compound 1 (10
mg)was obtained from C-5 (45 mg)by silica gel CC
eluted with petroleum ether /EtOAc (3∶ 1)．
3-Methyl-agallochaol C (1) colorless oil，C21 H34
O4，［α］
19
D -15． 49 (c 0． 57，CHCl3)． ESI-MS (neg．)
m/z 373 (61，［M + Na］+)．
Bioactivities
HIV-1NL4-3 Ｒeplication Inhibition Assay
A previously described HIV-1 infectivity assay was
used ［6，7］．
Cytotoxicity analysis
Cytotoxicity was determined by the sulforhodamine B
(SＲB)colorimetric assay ［8］．
Ｒesults and Discussion
Compound 1 was isolated as colorless oil． Its molecular
formula，C21 H34 O4，was determined by ESI-MS (m/z
373，［M + Na］+)，in combination with 1H and 13C
NMＲ data (Table 1)，indicating five degrees of unsat-
uration． In the 1H-NMＲ spectrum，a methoxyl group at
δH3． 63 (3H，s)，two tertiary methyls at δH 1． 76 and
Table 1 1H (600 MHz)and 13C (150 MHz)NMＲ data of
compound 1 in C5D5N
No． δH(mult．，J，Hz) δC(mult．)
1 1． 68 (2H，m) 34． 7 (t)
2 2． 37 (1H，overlap) 29． 4 (t)
2． 46 (1H，m)
3 － 175． 0 (s)
4 － 148． 5 (s)
5 2． 02 (1H，dd，J = 2． 6，12． 8) 51． 4 (d)
6 1． 27 (2H，m) 25． 6 (t)
7 1． 16 (1H，m) 39． 1 (t)
1． 28 (1H，m)
8 － 33． 7 (s)
9 1． 63 (1H，m) 44． 0 (d)
10 － 40． 4 (s)
11 1． 22 (1H，m) 24． 1 (t)
2． 39 (1H，overlap)
12 2． 25 (1H，m) 33． 4 (d)
13 1． 84 (1H，overlap) 24． 1 (t)
1． 44 (1H，m)
14 1． 84 (1H，overlap)0． 85 (1H，m) 28． 1 (t)
15 1． 37 (1H，d，J = 13． 6) 54． 1 (t)
1． 52 (1H，dd，J = 13． 6，2． 8)
16 － 74． 2 (s)
17 3． 85 (1H，d，J = 11． 0) 70． 0 (t)
3． 93 (1H，d，J = 11． 0)
8001 Nat Prod Ｒes Dev Vol. 27
18 1． 76 (3H，s) 24． 3 (q)
19 4． 80 (1H，br s) 114． 1 (t)
4． 93 (1H，br s)
20 0． 93 (3H，s) 18． 5 (q)
-OCH3 3． 63 (3H，s) 51． 9 (q)
0． 93 (each 3H，s)，two protons of terminal double
bond at δH 4． 80 and 4． 93 (each 1H，br s)，as well as
a pair of hydroxylmethyl protons at δH 3． 85 and 3． 93
(each 1H，d，J = 11． 0)were observed． Apart from
the methoxyl group (δC51． 9，q)，twenty carbons were
observed in the 13C NMＲ spectrum，including two
methyls，nine methenes (including an oxygenated one
and a terminal double bond)，three methines，four qua-
ternary carbons (containing an oxygenated one and an
olefinic one)and one carbonyl group． Comparison of
the 1H and 13C NMＲ spectroscopic data with those of
agallochaol C ［9］showed they were very similar，except
for the presence of the methoxyl group (δH 3． 63，s;δC
51. 9，q)in 1． This methoxyl was attached to C-3 (δC
175． 0，s)because it showed HMBC correlation with C-
3． Thus，the structure of 1 (Fig． 1)was determined and
named as 3-methyl-agallochaol C，which was a new nat-
ural product synthesized by Guo et al． in order to fur-
ther confirm the structure of agallochaol C ［9］．
Fig. 1 Chemical structures of compounds 1-14
The known compounds were determined to be eurifoloid
D (2)［5］，ent-3β，(13S)-dihydroxyatis-16-en-14-one
(3)［10］，ent-16α，17-Dihydroxyatisan-3-one (4)［10］，
ent-atisane-3β，16α，17-triol (5) ［11］，4，13β-di-
hydroxy-14-oxo-3，4-secoatis-16-en-3-oic acid methyl
ester (6)［12］，13β，19-dihydroxy-3，15-dioxoatis-16-
ene (7 )［12］， 13β-hydroxy-3， 15-dioxoatis-16-ene
(8)［12］，ent-16α，17-dihydroxykauran-3-one (9)［13］，
3-acetoxymethyl-5-［(E)-3-acetoxy-propen-1-yl) ］-2-
(4-hydroxy-3-methoxyphenyl)-7-methoxy-2，3-dihydro-
benzofuran (10)［14］，taraxerol (11)［15］，9β，19-cyclo-
lanostan-3β-ol (12)［15］，6，7，8-trimethoxyl-coumarin
9001
Vol. 27
LI Hong-mei，et al:Chemical Constituents from Euphorbia neriifolia and Their Ｒelated Anti-HIV and Anti-cancer Activities
(13)［15］，and 3，3-di-O-methylellagic acid (14)［15］
by comparison of their spectral data with literature val-
ues．
The anti-HIV-1 activity of compounds 3-5，8，9 and 11-
13 were evaluated using AZT as positive control (EC50
= 0． 0086 ± 0． 0015 μg /mL)，compounds 4 and 9
exhibited moderate anti-HIV-1 activity with EC50 values
of 6． 55 ± 2． 24 and 12． 3 ± 3． 75 μg /mL，respective-
ly． In addition，the cytotoxicities of compounds 3-5 and
8-13 against a panel of human cancer cell lines were
tested with Paclitaxel as positive control，and compound
10 possessed moderate anti-cancer activity (Table 2)．
Table 2 Cytotoxicity SＲB assay of compounds 3-5，8-13 and paclitaxel
Compounds
Cell Line (IC50，μM)
A549 MDA-MB-231 KB KB-VIN
3 ＞ 10 ＞ 10 ＞ 10 ＞ 10
4 ＞ 10 ＞ 10 ＞ 10 ＞ 10
5 ＞ 10 ＞ 10 ＞ 10 ＞ 10
8 ＞ 10 ＞ 10 ＞ 10 ＞ 10
9 ＞ 10 ＞ 10 ＞ 10 ＞ 10
10 7． 248 ± 0． 136 4． 796 ± 0． 044 4． 659 ± 0． 011 5． 519 ± 0． 107
11 ＞ 10 ＞ 10 ＞ 10 ＞ 10
12 ＞ 10 ＞ 10 ＞ 10 ＞ 10
13 ＞ 10 ＞ 10 ＞ 10 ＞ 10
Paclitaxel 0． 005230 ± 0． 000852 0． 004354 ± 0． 000978 0． 002899 ± 0． 000202 1． 298787 ± 0． 060698
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