全 文 ： 天然产物研究与开发
2005 Vol.17 No.6 NATURAL PRODUCT RESEARCH AND DEVELOPMENT
Received December 27 ,2004;Accepted January 31, 2005
＊Corresponding author Tel:86-21-54237564;E-mai l:cqhu@shmu.edu.
cn
Taxiphyllin:A Cyanogenic Glucoside with Tyrosinase Inhibitory
Activity from the Shoots of Pleioblastus amarus
CHENG Ke-jun1 ,CHEN Jing1 ,LIANG Gao-lin1 ,
YAO Hui2 ,LIN Hui-fen2 ,WEI Shao-min2 ,YANG Jun1 ,HU Chang-qi1＊
(1.School of Pharmacy , Fudan University , Shanghai 200032 , China;
2.Shanghai Jahwa United Co., Ltd ., Shanghai 200056 , China)
Abstract:A cyanogenic glycoside , taxiphyllin(1), was isolated from the shoots of Pleioblastus amarus(Keng)Keng f.for the
first time.Its structure was elucidated on the basis of spectroscopic analysis(UV , IR,MS , 1D and 2D NMR)and physiochemi-
cal properties.Taxiphyllin was found to inhibit tyrosinase activity in vitro significantly and considered to be a potent tyrosinase
inhibitor.
Key words:Pleioblastus amarus;taxiphyllin;cyanogenic glucoside;ty rosinase
Taxiphyllin:苦竹笋中具有酪氨酸酶抑制活性的氰苷
程科军1 ,陈 竞1 ,梁高林1 ,姚 慧2 ,林惠芬2 ,魏少敏2 ,杨 君1 ,胡昌奇1＊
(1.复旦大学药学院 , 上海 200032;2.上海家化联合股份有限公司 ,上海 200056)
摘 要:从苦竹笋 Pleioblastus amarus(Keng)Keng f.中首次分得一个氰苷化合物 , 通过光谱分析及其物理化学性
质鉴定为 taxiphyllin(1)。该化合物在体外能显著抑制酪氨酸酶活性 ,是一个强有效的酪氨酸酶抑制剂。
关键词:苦竹笋;taxiphyllin;氰苷;酪氨酸酶
中图分类号:R284.1;Q946.91
Introduction
Pleioblastus amarus(Keng)Keng f.(Gramineae)occurs
widely in south China.The shoots of the plant are used in
Chinese herbal medicine as antipyretic and diuretic a-
gents[ 1] .In the southwest of China , there is a long history
that women use them as brightening agents by means of
washing faces with the water in which the shoots of the
plant are marinated.Previous chemical work on the plant
resulted in the isolation of several flavonoids and other
compounds from the leaves of the plant
[ 2-5] .In the screen-
ing of bioactivity , the shoots of Pleioblastus amarus
(Keng)Keng f.displayed a significant inhibitory activity
against tyrosinase in vitro .For this reason ,we systemati-
cally investigated the chemical constituents of the shoots of
the plant and isolated a cyanogenic glucoside in a sizable
quantity from the n-BuOH extraction.The present paper
deals with the isolation , structure elucidation and in vitro
tyrosinase inhibitory activity of the cyanogenic glucoside ,
taxiphyllin (1),which was obtained from the plant for the
first time.
Results and Discussion
Identification of compound 1
Compound 1 was obtained as white needles and was as-
signed the molecular formula of C14H17NO7 based on the
element analysis , ESI-MS ,HR EI-MS , 13C NMR and 1H
NMR data.The element analysis indicated that 1 has a ni-
trogen element.The peak 2250 cm-1 in IR and the signal
δ118.54 in 13C NMR suggested the existence of a nitrile
group.The 1H NMR spectrum(400MHz ,DMSO-d6)ex-
hibited the typical AA BB pattern (δ7.32 and δ
6.82 , J =8.6Hz ,4H)of a p-substituted aromatic ring , a
sharp single at δ5.87(1H , methane-H), a poorly re-
solved doublet of the anomeric glucose proton (δ4.07 ,
1H , J =7.02 Hz , β-configuration), the remaining sugar
protons (δ3.68 , δ3.47 ,2H and δ3.05 ,4H )and the
733
hydroxyl proton (δ9.84 , δ5.23 , δ5.05 , δ4.98 and δ
4.55 , 5H ).The 1H NMR data were consistent with the
published data for the cyanogenic glucoside taxiphyllin[ 6] .
The 13C NMR data presented in Table 1 were obtained for
the first time in DMSO-d6 and are similar to values with
acetone-d6 as the solvent in which the cartons are almost
shifted downfield in 1 ～ 2 ppm respectively[ 7] .The HSQC
and HMBC confirmed the values of each carbon .The peak
at m/z 311 in HR EI-MS is the molecular ion and the
data at m/z 334 [M+23] + ,350 [M+39] + in ESI-MS
coincides with the estimation of it.TheMS also show typi-
cal glucose peaks at m/z 60 , 61 ,73 , 91 ,127 , 145 ,163.
There were two alternative ways of breaking the glucosidic
bond ,which give peaks at m/z 149 and 132.All these
MS data were identical with the data of taxiphyllin in the
literature[ 8] .On the basis of the above evidences , 1 was
elucidated as taxiphyllin ,2R-β-D-glucopyranosyloxy-2-(4-
hydroxy)-phenylacetonitrile.
Taxiphyllin and the plants
Before Pleioblastus amarus(Keng)Keng f., taxiphyllin
had been found in Taxus canadensis(Taxaceae), Taxus
baccata (Taxaceae), Triglochin maritima (Juncagi-
naceae), Girgensohnia oppositiflora (Chenopodiaceae),
Henriettella fascicularis(Melastomataceae)and some bam-
boo shoots mainly including Dendrocalamus latiflorus ,
Dendrocalamus giganteus , Dendrocalamus hamiltonii ,
Bambusa vulgaris , Bambusa guadua [ 7 ,9-14] .It has been
known for a long time that some species of bamboo from
which bamboo shoots are derived can contain cyanide- or
more properly a cyanogenic glucoside.Taxiphyllin was the
cyanogenic glucoside in some kinds of bamboo shoots and
it was an instable component especially when placed un-
der the acid condition .In the process of isolating taxi-
phyllin ,we discovered the existence of a sizable quantity
of p-hydroxybenzaldehyde(2), which may be partly hy-
drolyzed from taxiphyllin by the enzymes in the plant or
some chemical reagents (such as CHCl3)used as the elu-
ent.
Taxiphyllin , a bitter principle , sometimes makes bamboo
shoots too bitter to eat and potentially toxic.Nahrstedt[ 15]
found that the compound would decomposes quickly when
placed in boiling water.Ferreira et al[ 16] noted that boil-
ing bamboo shoots for 20 min at 98 ℃ removed nearly
70% of the HCN while all improvements on that (higher
temperatures and longer intervals)removed progressively
up to 96%, thus even the highest quoted figure(800 mg/
100 g)of taxiphyllin would be de-toxified by cooking for
two hours.Because of this instability , bamboo shoots be-
comes edible after proper cooking.
Tyrosinase inhibitory activity of taxiphyllin
Tyrosinase(monophenol monooxygenase;E.C .1.14.18.
1)also known as polyphenol oxidase(PPO), is a copper-
containing monooxygenase widely distributed in nature.It
is responsible for the synthesis of melanin in animals and
browning in plants.A recent review gave a comprehensive
understanding of it
[ 17] .Although a large number of natu-
rally occurring tyrosinase inhibitors have already been de-
scribed[ 18] , their individual activity is ether not potent e-
nough to be considered of practical use or safety regula-
tions concerning food additives limit their in vivo use.
Therefore ,more potent compounds are needed .
Taxiphyllin have been tested for the tyrosinase inhibitory
capacity and present primary data suggest it a potential ty-
rosinase inhibitor.It is of interest to find L-tyrosine(3)is
the precursor of taxiphyllin
[ 19]
in many plants and L-tyro-
sine is just the substrates of the tyrosinase.Due to the
close structural similarity of 1 , 2 and 3 , it is considered
that 1 and 2would be classical competitive inhibitors.The
close analogues of 2 , p-substituted benzaldehydes , have
been classified as competitive inhibitors and the inhibitory
activity has been attributed to the existence of the alde-
hyde group[ 20] .The further studies on the mechanisms of
tyrosinase inhibiting of taxiphyllin and the toxicity of taxi-
phyllin are under investigation.
Experimental
General
UV spectra were obtained in absolute MeOH on a Shi-
madzu UV-260 spectrophotometer.IR spectra were mea-
sured in KBr pellets on a Nicolet Avatar 360 E.S .P.
Fourier Transform Infrared Spectroscopy.ESI-MS and HR
EI-MS were taken on a Q-Tof micro spectrometer.NMR
spectra (1H NMR , 13C NMR , HSQC and HMBC)were
recorded in DMSO-d6 on a Mercury Plus 400 NMR spec-
trometer using TMS as internal standard .Chemical shift
vaules were reported in δ(ppm)units and referenced to
DMSO-d6 at δH 2.49 and δC 39.5 respectively.Coupling
constant J were expressed in Hz.Macroporous absorption
resin DIAION HP-20 was obtained from Mitsubishi Chem-
ical Corporation (Japan)and silica gel was supplied by
Qingdao Marine Chemical Ltd.,China.Tyrosinase (E .C.
1.14.18.1), L-DOPA(L-3 , 4-dihydroxyphenylalanin),
734 天然产物研究与开发 2005 Vol.17 No.6
arbutin and kojic acid(5-hydroxy-2-hydroxymethyl-4H-
one)were purchased from Sigma-Aldrich Inc..
Plant material
The shoots of Pleioblastus amarus(Keng)Keng f .were
collected from the Wanzhou District of Chongqing Munici-
pality in China and the bamboo was identified by Prof.
Pan Sheng-li ,School of Pharmacy ,Fudan University.
Extraction and isolation
The shoots of Pleioblastus amarus (Keng)Keng f.(55
kg)were extracted with 75% ethanol at room tempera-
ture.The extract was concentrated under reduced pressure
to afford 882 g of residues , which were suspended with
water.The suspended aqueous solution was partitioned
with petroleum ether , chloroform , ethyl acetate and n-
BuOH successively.The n-BuOH extract were evaporated
to give 103.3 g of residues ,which were subjected to chro-
matography over macroporous absorption resin DIAION
HP-20 eluted with water and different concentrations of
ethanol.Further chromatography of 10% ethanol extract
over silica gel (200 ～ 300 mesh)with chloroform-
methanol(10∶1 ～ 1∶1)as the mobile phase yielded the
compound 1(1.81 g).
Bioactivity test
Tyrosinase inhibitory activity was tested in Shanghai Jahwa
United Co.,Ltd.and the assay method was similar to the
literature[ 21 , 22] .The assay procedure was based on the
spectrophotometric detection of dopachrome at an ab-
sorbance of 475 nm , formed by the oxidation of L-DOPA
by tyrosinase as originally described by Mason
[ 23] .It was
found that 1 can inhibit the oxidation of L-DOPA with an
IC50 of 32 μM.Kojic acid and arbutin were used as posi-
tive standards with an IC50 of 35 μM and 327 μM respec-
tively.The data strongly suggest that 1 would be a highly
effective tyrosinase inhibitor.
Physical Costant and Spectrum Data
Taxiphyllin , white needles (from MeOH).UV(MeOH)
λmax nm:231.8 , 274.8.IR (KBr)νmax cm-1 :3600 ～
2400 ,1070 , 1050 , 1008 , 896 (β-glucosyl);1617 , 1521 ,
825(p-substituted aromatic ring);2250(nitrile).HREI-
MS:311 , 163 , 161 , 149 , 145 , 134 , 133 , 132 , 131 , 127 ,
103 ,91 , 73 , 61 ,60;ESI-MS (partly ,not pure samples):
350 [ M +39] + , 334 [ M +23] + , 132.1H NMR(400
MHz ,DMSO-d6)δ:9.84(1H , s ,OH-4),7.32(2H , d , J
=8.6 Hz ,H-2 , 6), 6.82(2H , d , J =8.6 Hz ,H-3 , 5),
5.87(1H , s , H-7), 5.23(1H , s , OH-2′), 5.05(1H , s ,
OH-4′),4.98(1H , s ,OH-3′),4.55(1H , t ,OH-6′), 4.07
(1H ,d , J =7.02 Hz ,H-1′), 3.68(1H , q ,H-6a), 3.45
(after D2O addition ,1H ,dd , J 1=11.3 Hz , J 2 =3.9Hz ,
H-6b),3.04(4H , s ,H-2′, 3′,4′,5′).13C NMR data see
Table 1.
Table 1 The 13 C NMR spectral data of compound 1(100
MHz , in DMSO-d6)
C 1 1＊
1 118.5 119.6
2 65.9 67.8
3 123.0 125.3
4 128.9 130.4
5 115.2 116.6
6 157.8 159.8
7 115.2 116.6
8 128.9 130.4
1′ 99.7 101.5
2′ 72.9 74.6
3′ 76.3 78.0
4′ 69.8 71.7
5′ 77.1 78.0
6′ 61.1 63.1
＊Data f rom the literature measured in acetone-d 6
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