全 文 ：担子菌地星菌丝体中一个新的甾醇酯
?
邵红军1 , 2 , 房立真1 ,2 , 杨婉秋1 , 2 , 王 飞1 , 2 , 刘吉开1
??
(1 中国科学院昆明植物研究所 植物化学与西部植物资源持续利用国家重点实验室 , 云南 昆明 650204;
2 中国科学院研究生院 , 北京 100049 )
摘要 : 从担子菌地星 ( Astraeus hygrometricus) 发酵培养菌丝体中分离得到一个新的多羟基甾醇酯 , 其化学
结构通过波谱学方法包括二维核磁共振鉴定为 : 3β, 5α-二羟基- (22 E , 24 R ) -麦角甾醇-7 , 22-二烯-6α-棕榈
酸酯。同时还从该菌中分离得到其它三个甾醇类化合物。
关键词 : 地星 ; 甾醇酯 ; 担子菌
中图分类号 : Q 946 文献标识码 : A 文章编号 : 0253 - 2700 (2007) 03 - 371 - 04
A New Steryl Ester from the Culture Mycelia of the
Basidiomycete Astraeus hygrometricus (Astraceae)
SHAO Hong-J un
1 , 2
, FANG Li-Zhen
1 , 2
, YANG Wan-Qiu
1 , 2
, WANG Fei
1 , 2
, LIU Ji-Kai
1 **
( 1 State Key Laboratory of Phytochemistry and Plant Resources in West China , Kunming Instituteof Botany,
Chinese Academy of Sciences, Kunming 650204 , China;
2 GraduateSchool of Chinese Academy of Sciences, Beijing 100049 , China)
Abstract : A new steryl esterwith a polyhydroxylated ergostane-type nucleus, 3β, 5α-dihydroxy- (22 E , 24 R ) -ergosta-7 ,
22-dien-6α-yl palmitate (1) , togetherwith threeknowncompounds (2 - 4 ) was isolatedfromthe culturemyceliaof theba-
sidiomycete Astraeus hygrometricus . The structure of compound 1 was elucidated on the basis of extensive spectroscopic
methods ( IR , HR-FAB-MS, 1D and 2D NMR) .
Key words: Astraeus hygrometricus; Steryl ester; Basidiomycete
Astraeus hygrometricus (Sclerodermatales, Basid-
iomycete) , a mycorrhizal fungus, is widely distributed
in China . It is also used as a remedy for haemastatic
and inflammation in traditional Chinese medicine
(Mao, 2000 ) . Three triterpenes and a splenocyte ac-
tivity glucan have previously been isolated from the
fruiting bodies of this fungus ( Takaishi et al . 1987;
Chakrabory et al . 2004 ) . To the best of our knowl-
edge, there are no chemical investigations on the cul-
ture broth of this fungus . In our continuing studies on
thebasidiomycete-derived secondary metabolites (Liu,
2002 , 2005 , 2006; Shao et al . 2005; Wang and Liu,
2005) , we have been isolated anew steryl ester with a
polyhydroxylated ergostane-type nucleus, 3β, 5α-di-
hydroxy -( 22 E , 24 R )-ergosta-7 , 22-dien-6α-yl palmi-
tate (1) (Fig . 1 ) , as well as three known compounds
( 22 E, 24 R ) -5α, 8α-epidioxyergosta-3 , 22-dien-3β-
ol ( 2) , ( 22 E, 24 R )-ergosta-4 , 6 , 8 (14 ) , 22-tet-
raen-3β-one ( 3 ) , and ( 22 E , 24 R )-ergosta-7 , 22-
dien-3β-ol (4 ) fromthe culturemyceliaof the fungus .
It is noted that there are few naturally occurring poly-
hydroxylated steryl esters reported up to date ( Wang
and Liu, 2005; Yang et al . 2003; Zhang et al .
2005) . This paper deals with the isolation and struc-
云 南 植 物 研 究 2007 , 29 (3) : 371～374
Acta Botanica Yunnanica

?
?? ?Corresponding author . Tel : + 86 - 871 - 5216327; E-mail : jkliu@ mail .kib. ac. cn
Received date: 2006 - 12 - 05 , Accepted date: 2007 - 02 - 04
作者简介 : 邵红军 (1977 - ) 男 , 博士研究生 , 主要从事高等真菌化学和活性研究。 ?
Foundation item: This work was supported by theNational Natural Science Foundation of China ( 30470027 and 30225048 ) and theNatural Science
Foundation of Yunnan Province ( 2005C0011R and 2005C0052M ) .
ture elucidationof the new steryl ester (1) .
Results and Discussion
Compound 1 was obtained as a colorless oily sol-
id . Themolecular formulawas determined to be C44 H76
O4 by
13 C-NMR data and HR-FAB-MS ( calc . for [M
+ H] + : 669 .5821; found: 669 .5814) . Its IR spec-
trumrevealed the presence of hydroxyl groups ( 3 431
cm
- 1 ) . The 1 H-NMR spectrum ( Table 1 ) which as-
signed with aid of the
1
H-
1
H COSY spectrum ( Fig .
2) , exhibited two tertiary methyl signals atδ1 .01 (s,
H - 19) and 0 .56 (s, H - 18 ) , four secondary methyl
signals atδ1 .02 ( d, J = 6 .6 , H - 21) , 0 .91 ( d, J
= 6 .9 , H - 28 ) , 0 . 83 ( d, J = 7 .2 , H - 27 ) and
0 .82 ( d, J = 7 .2 , H - 26 ) , a pair of 1 , 2-disubsti-
tutedolefinic protons atδ5 .22 ( dd, J = 15 .2 , 7 .4 , H
- 23) and 5 .15 ( dd, J = 15 .2 , 8 .0 , H - 22) , and a
trisubstitutedolefinic proton atδ4 .88 ( s, H - 7 ) . Its
13
C-NMR analyzed together with the DEPT and HMQC
spectra also showed signals due to an oxygenated qua-
ternary carbon atδ75 .2 ( s, C - 5 ) , two oxygenated
methane carbons atδ73 .7 ( d, C - 6 ) and 67 .3 ( d, C
- 3) , and four olefinic carbons atδ144 .2 (s, C - 8) ,
135 .4 ( d, C - 22 ) , 132 .2 ( d, C - 23 ) and 115 .2
( d, C - 7 ) . Based on these data, 1 was suggested to
be an ergosta-7 , 22-dien-3β, 5α, 6-triol derivatives .
Meanwhile in
1
H-NMR spectrum, a characteristic
downfield signal of H - 6 (δ5 .27 , s) caused by acyla-
tion effect distinctly indicated that thepalmitatemoiety
was located at 6αposition of the sterol nucleus . This
was also supportedby a cross peak observed between H
- 19 and H - 6β in the ROSEY spectrum . These data
were in good agreement with those reported for ergosta-
7 , 22-dien-3β, 5α, 6α-triol and its derivatives (Ayer
et al . 1992; Chen et al . 1991; Goldstein and Frye,
1996) . Furthermore, the 1 H- and 13 C-NMR of com-
pound 1 revealed a terminal methyl signal at (δ0 .88
( t, J = 6 .8 , H - 16′)?14 .1 ( q, C - 6′) ) , a methyl-
ene group inα-position to an ester function at (δ2 .30
( t, J = 7 .6 , H - 2′)?34 .6 ( t, C - 2′) ) , a carbonyl
carbon atδ173 .4 ( s, C - 1′) , and other signals at
(δ1 .64 (m, H - 3′) , 1 . 25 - 1 .34 (overlapped) , and
δ31 .4 ( t, C - 14′) , 25 .1 ( t, C - 3′) , 22 .7 ( t, C -
15′) , 29 .2 - 29 .8 ( t) ) , which showed to be a saturat-
ed long-chain fatty-acid ester moiety . By comparison
with the data in literature (Zhang et al . 2005 ) , those
data indicated that the saturated long-chain fatty-acid
was palmitate . It was also proved by the EI-MS data,
which displayed fragment ion peaks at m?z 412 ( 67
[M-C16 H32 O2 ] + ) , 394 ( 34 [M-C16 H32 O2 -H2 O] + )
and 376 (27 [M-C16 H32 O2 -2H2 O] + ) . The linked po-
sition of the palmitate moiety was further confirmed by
the clearly correlation between H - 6β (δ5 .27 , s) and
C - 1′(δ173 .4 , s) in the HMBC spectrum ( Fig . 2 ) .
The geometry of the △22 -double bond was determined
to be E from the coupling constant ( J = 15 .2 ) be-
tween H - 22 and H - 23 . The stereochemistry at C - 20
and C - 24 was deduced to be R and R, respectively,
by comparison of
1
H- and
13
C-NMR data with those of
ergosterol (Wright et al . 1978) . Fromall abovedata,
the structure of 1 was assigned as 3β, 5α-dihydroxy -
(22 E , 24 R )-ergosta-7 , 22- dien-6α-yl palmitate .
Table 1 1 H- and 13 C-NMR dataof Compound 1 ( CDCl3 ) .
Position δC δH Position δC δH
1 Z31 ?. 9 ( t) 1 . 25 ( m) 19 y17 .9 ( q) 1 }. 01 ( s)
2 Z30 ?. 8 ( t) 1 . 85 ( m) 20 y40 .3 ( d) 2 }. 02 ( m)
3 Z67 y. 3 ( d) 4 ?. 00 ( m) 21 21 .1 ( q) 1 }. 02 ( d, J = 6 C. 6)
4 Z39 ?. 2 ( t) 2 . 04 ( m) , 1 ]. 92 ( m) 22 y135 .4 ( d) 5 }. 15 ( dd, J = 15 .2 , 8 @. 0 )
5 Z75 }. 2 (s) - 23 y132 ?. 2 ( d) 5 . 22 ( dd, J = 15 .2 , 7 @. 4 )
6 Z73 y. 7 ( d) 5 ?. 27 (s) 24 42 .9 ( d) 1 }. 85 ( m)
7 Z115 ?. 2 ( d) 4 . 88 (s) 25 y33 .1 ( d) 1 }. 47 ( m)
8 Z144 ?. 2 (s) - 26 y19 .6 ( q) 0 }. 82 ( d, J = 7 C. 2)
9 Z43 y. 5 ( d) 2 ?. 09 ( m) 27 19 .9 ( q) 0 }. 83 ( d, J = 7 C. 2)
10 l39 }. 0 (s) - 28 y17 ?. 6 ( q) 0 . 91 ( d, J = 6 C. 9)
11 l21 ?. 3 ( t) 1 . 57 ( m) 1 Y′ 173 ?. 4 (s) -
12 l39 ?. 6 ( t) 1 . 52 ( m) , 1 ]. 30 ( m) 2 Y′ 34 .6 ( t) 2 }. 36 ( t, J = 7 3. 6)
13 l43 }. 8 (s) - 3 Y′ 25 ?. 1 ( t) 1 . 64 ( m)
14 l54 y. 8 ( d) 1 ?. 90 ( m) 4 ?′- 13′ 29 .2 - 29 X. 8 ( t) 1 }. 25 - 1 E. 34 ( m)
15 l22 ?. 7 ( t) 1 . 40 ( m) 14 k′ 31 .4 ( t) 1 }. 25 - 1 E. 34 ( m)
16 l28 ?. 0 ( t) 1 . 73 ( m) 15 k′ 22 .7 ( t) 1 }. 25 - 1 E. 34 ( m)
17 l55 y. 9 ( d) 1 ?. 28 ( m) 16 k′ 14 .1 ( q) 0 }. 88 ( t, J = 6 3. 8)
18 l12 y. 2 ( q) 0 ?. 56 (s)
Assignment made on thebasis of 1 H-1 H COSY , HMQC and HMBC data .
273 云 南 植 物 研 究 29 卷
Fig . 1 The structureof compound 1 Fig . 2 1 H-1 H COSY and key HMBC correlations of compound 1
Compounds2, 3, and 4 werealso isolated fromthe
same fungus, and identified as ( 22 E, 24 R)-5α, 8α-
epidioxyergosta-3 , 22-dien-3β-ol ( 2 ) , ( 22 E, 24 R )-
ergosta-4, 6, 8 ( 14 ) , 22-tetraen-3β-one ( 3 ) , and
( 22 E, 24 R )-ergosta-7 , 22-dien-3β-ol ( 4 ) , respec-
tively, according to the physical and spectroscopic data
in literatures (Yue et al . 2001; Gao et al . 2001; Lu
et al . 1985) .
Experimental
General Optical rotation was measured on a Horiba SE-
PA-300 spectropolarimeter . IR spectrum was obtained with a
Bruker Tensor 27 spectrometer, with KBr pellets, in cm- 1 . 1D
and2D-NMR spectrawere recorded on Bruker AV-400 and DRX-
500 spectrometers in CDCl3 , δ in ppm, J in Hz . EI-MS was
recorded with a Thermo Finnigan Trace DSQ spectrometer , HR-
FAB-MS was recorded with a VG Autospec-3000 spectrometer .
Silica gel (200 - 300 mesh, Qingdao Marine Chemical Inc,
Qingdao . P . R . China) , and Sephadex LH-20 (Amersham Bio-
sciences) were used for column chromatography . Pre-coated sili-
ca gel GF254 plates (Qingdao Marine Chemical Inc, Qingdao . P .
R . China) wereused for TLC . Fractionsweremonitored by TLC
and spotswerevisualized by heating silicagel plates sprayed with
10% H2 SO4 in EtOH .
Mushroom Material and Culture The fungus A. hygr-
ometricuswas collected at the Botanic Garden of Kunming Insti-
tute of Botany, Chinese Academy of Sciences, P . R . China, in
September 2001 , and identified by Dr . Fu-Qiang Yu, Kunming
Institute of Botany . The voucher specimen was deposited in the
Herbariumof Kunming Institute of Botany, Chinese Academy of
Sciences . Culture medium: potato ( peeled) 200 g, glucose 20
g, KH2 PO4 3 g, MgSO4 1 .5 g, citric acid 0 .1 g and thiamin
hydrochloride10 mgin 1 L of deionized H2 O ( pH 6.5 beforeau-
toclaving) . The culture liquid was fermented at 25℃ for 20 days
on a rotary shaker (150 r?min) .
Extraction and Isolation The dried mycelia (106 g) fil-
tered fromculture broth (25 L ) were successively extracted with
CHCl3?MeOH (1∶1) . The extract was evaporated in vacuo and
theoily residue (8 . 0 g) was subjected to CC (silica gel , petro-
leumether?Me2 CO (9∶1→1∶1) ) . The fraction (0 . 36 g) from
petroleum ether?Me2 CO (20∶1) was further purified byCC (sili-
ca gel, petroleum ether?AcOEt 20∶1 ; Sephadex LH-20 , CHCl3?
MeOH 1∶1 ) to afford the pure compound 3 ( 10 mg) and 4 ( 13
mg) . Thefraction (0 . 51 g) from petroleum ether?Me2 CO ( 10∶
1) was further isolated by CC (silica gel , CHCl3?AcOEt 20∶1)
to give the pure compound1 (6 mg) and 2 (20 mg) .
Compound 1 , C44 H76 O4 , colorless oily soild; [α]24.4D = + 46
( c 0 .17 , CHCl3 ) ; IRνKBrmax cm- 1 : 3431 , 2956 , 2925 , 2852 ,
1710 , 1629 , 1461 , 1381 , 1187; 1 H- and13 C-NMR data: seeTa-
ble 1; EI-MS: 412 (67 [M-C16 H32 O2 ] + ) , 394 (34 [M-C16 H32
O2 -H2 O] + ) , 376 (27 [M-C16 H32 O2 -2H2 O] + ) , 251 (100) , 157
(48) , 129 (30) , 109 (15 ) , 69 (56 ) ; HR-FAB-MS ( pos .) :
669 .5814 ( [M+ H] + , C44 H77 O4 + , calc . 669 .5821) .
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第五届世界菌根食用菌大会消息
由云南楚雄彝族自治州人民政府 , 中国食品土畜进出口商会 , 中国菌物学会 , 中国科学院昆明植
物研究所主办 , 云南省林业厅 , 云南省商务厅等 21 家共同协办的第五届世界菌根食用菌大会将于
2007 年 8 月 26～29 日云南楚雄召开。会议将由菌根食用菌的科技论坛 , 商业贸易论坛和文化论坛
(野生食用菌科普展览和云南野生食用菌品尝 ) 三部分组成。
大会组委会
主席 : 王云 副主席 : 杨红卫、曹绪岷、李玉、刘培贵 委员 : 延荣科、杨胜军、郭辉军、张世忠
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有关大会详细信息可登陆网站 www. iwemm5. com 查询
The 5th International Workshop of Edible Mycorrhizal Mushrooms Organized by People′s Government
of Chuxiong Yi Nationality Autonomous prefectureof Yunnan Province, China Chamber of Commerce of Import and
Export of Foodstuffs NativeProduce and Animal By-Products, Mycological Society of China, Kunming Instituteof Bot-
any, Chinese Academy of Sciences, Kunming, China, the Fifth International Workshop on Edible Ectomycorrhizal
Mushrooms ( IWEMM5) will be held in Chuxiong, Yunnan, China from26 to 29th August 2007 . All aspects of the
scienceof edible ectomycorrhizal mushrooms will be covered in the workshop . Considerable time will also be devoted
to the tradeand cultivationof EEMM whichweanticipatewill beof benefit to the economic development of the region .
Organizing Committee:
Chairman: Wang Yun, Vice-Chairman: Yang Hong-wei , Cao Xu-Min, Li Yu, Liu Pei-Gui
Members: Yan Rong-Ke, Yang Sheng-Jun, Guo Hui-Jun, Zhang Shi-Zhong
International Scientific Committee:
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Members: ?Shannon Berch, Canada, Gérard Chevalier, France, Eric Danell , Sweden rance, Eric Danell , Sweden Eric Danell,
Sweden rance, Eric Danell, Sweden
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473 云 南 植 物 研 究 29 卷