Genus Caulophyllum: An Overview of Chemistry and Bioactivity

Abstract

Recently, some promising advances have been achieved in understanding the chemistry, pharmacology, and action mechanisms of constituents from genus Caulophyllum. Despite this, there is to date no systematic review of those of genus Caulophyllum. This review covers naturally occurring alkaloids and saponins and those resulting from synthetic novel taspine derivatives. The paper further discussed several aspects of this genus, including pharmacological properties, mechanisms of action, pharmacokinetics, and cell membrane chromatography for activity screening. The aim of this paper is to provide a point of reference for pharmaceutical researchers to develop new drugs from constituents of Caulophyllum plants.

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Review Article
Genus Caulophyllum: An Overview of Chemistry and Bioactivity
Yong-Gang Xia,1Guo-Yu Li,2Jun Liang,1Bing-You Yang,1
Shao-Wa Lü,1and Hai-Xue Kuang1
1Key Laboratory of Chinese Materia Medica, Heilongjiang University of Chinese Medicine,
Ministry of Education, Harbin 150040, China
2Pharmaceutical College, Harbin Medical University, Harbin 150086, China
Correspondence should be addressed to Hai-Xue Kuang; hxkuang@hotmail.com
Received  January ; Accepted  April ; Published  May 
Academic Editor: Yong Jiang
Copyright ©  Yong-Gang Xia et al. is is an open access article distributed under the Creative Commons Attribution License,
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Recently, some promising advances have been achieved in understanding the chemistry, pharmacology, and action mechanisms
of constituents from genus Caulophyllum. Despite this, there is to date no systematic review of those of genus Caulophyllum.is
review covers naturally occurring alkaloids and saponins and those resulting from synthetic novel taspine derivatives. e paper
further discussed several aspects of this genus, including pharmacological properties, mechanisms of action, pharmacokinetics, and
cell membrane chromatography for activity screening. e aim of this paper is to provide a point of reference for pharmaceutical
researchers to develop new drugs from constituents of Caulophyllum plants.
1. Introduction
Caulophyllum isasmallgenusofperennialherbsinthefamily
Berberidaceae. e genus Caulophyllum is well known for its
diversity and pharmacological uses in traditional medicine
system since ancient times. All species in this genus are very
similar []. C. robustum is native to eastern Asia, especially in
China, while C. thalictroides and C. giganteum are native to
eastern North America. It is worth noting that nearly all
phytochemical and pharmacological studies on this genus are
focused on C. thalictroides and C. robustum duetotheir
important medical functions [].
e roots and rhizomes of C. thalictroides (L.) Michx.
(blue cohosh) have been used traditionally by Native Ameri-
cans for medicinal purposes []. e primary function of blue
cohosh in many native communities of North America was to
induce childbirth, ease the pain of labor, rectify delayed or
irregular menstruation, and alleviate heavy bleeding and pain
during menstruation []. Between  and , blue cohosh
was listed in the United States Pharmacopoeia as a labor indu-
cer [] and sold as an herbal supplement that can aid in
childbirth. Dietary supplements of blue cohosh are readily
available throughout the USA over-the-counter and from
Internet suppliers []. ere is considerable concern about
the safety of blue cohosh with reports of new born babies hav-
ing heart attacks or strokes aer the maternal consumption
of blue cohosh to induce labor [–]. ere is a heated dis-
cussion about using blue cohosh as dietary supplements for
women [].
C. robustum Maxim is well-known in Hong Mao Qi in
Chinese, which grows widely throughout north-east, north-
west, and south-west China. Its roots and rhizomes have been
used as folk medicine to treat external injuries, irregular-
menses, and stomach-ache due to its strong and wide bio-
logical activities []. Modern pharmacological studies have
demonstrated that alkaloids and triterpence saponins are
responsible for its major biological function as an anti-
inammatory [], analgesic [], antioxidant [], antibacte-
rial [], antiacetylcholinesterase [], and antitumor [,].
Taspine, a lead compound in anticancer agent development
[,], was rstly screened to possess obvious eect on
tumor angiogenesis and human epidermal growth factor rec-
eptor by using cell membrane chromatography from the C.
robustum [].
So it is very necessary to deeply explore Caulophyllum
plants. In the past decades, some promising advances have
been achieved in understanding the chemistry, pharmacol-
ogy, and action mechanisms of constituents from genus
Hindawi Publishing Corporation
Evidence-Based Complementary and Alternative Medicine
Volume 2014, Article ID 684508, 18 pages
http://dx.doi.org/10.1155/2014/684508

 Evidence-Based Complementary and Alternative Medicine
Macroporous or (and) ion
exchange resins
Vacuum evaporated
Adjust pH 7.0,
Suspension in
pH 9.0 with
Organic layer
Vacuum evaporated
Vacuum
evaporated
Vacuum evaporated Adjust pH 7.0,
Adjust pH 9.0
Maceration or reux using methanol
or 70% ethanol for extraction
Total extract
Liquid-liquid extraction Column chromatography
Water layer
Water layer
Water layer
Organic layer
Total alkaloidal fraction
Saponin fraction
Neutral fraction
Specic elution
Saponin fraction
Weak base Fr. 1
Nonphenolic Fr. 2 Phenolic Fr. 3
Note: ①5% or 0.1 N HCl in H2O; ②H2O
CHCl3
CHCl3
1N NaOH
CHCl3or EtOAc
CHCl3or EtOAc
Suspension in ①
Suspension in ① or ②,
As followeds ① is mentioned
CHCl3or EtOAc
n-BuOH
Caulophyllum plant materials
n-BuOH layer
CHCl3residue
NH4OH, CHCl3
S : Summary of procedures for isolation of alkaloids and saponins from Caulophyllum plants.
Caulophyllum. From the opinion of safety of using dietary
supplements of blue cohosh, a review dealing with quan-
titative methods of primary constituents of blue cohosh in
dietary supplements has been published []. However, to date,
thereisnosystematicreviewofchemistry,pharmacology,and
action mechanisms of constituents from genus Caulophyllum.
In this review, the dierent structures of the alkaloids
and saponins in genus Caulophyllum are described, including
naturally occurring constituents and synthetical taspine
derivatives. e present review highlighted the chemistry and
pharmacological diversity and mechanism of action. e aim
of this paper is to provide a point of reference on Caulo-
phyllum plants for pharmaceutical researchers. Furthermore,
various perspectives and existing problems for this genus are
oered for consideration.
2. Phytochemistry
Phytochemical research carried out on genus Caulophyllum
ledtotheisolationofalkaloidsandtriterpencesaponinsanda
few other classes of secondary metabolites. A comprehensive
summary of structures and isolation methods of metabolites
classied by structural types was given in present review.
Scheme summarizes the procedures for crude isolation of
alkaloids and triterpene saponins from genus Caulophyllum.
e roots and rhizomes of Caulophyllum plants are extracted
with methanol or % ethanol by maceration [,]orreux
[], and the combined extracts are concentrated in vacuo to
dryness. en two schemes are available for acquiring the
alkaloid and saponin fractions, namely, liquid-liquid parti-
tion and liquid-solid column chromatography methods [].
Liquid-liquid partition is commonly performed for crude
isolation. In most cases, the residue is suspended in % or
.NHClinwaterandthenpartitionedwithEtOAcor
CHCl3to remove neutral constituents. e aqueous layer was
then removed, NH4OHwasaddedtomakeitbasic(pH),
andthewholewasextractedwithEtOAcorCHCl
3.e
EtOAc or CHCl3soluble part was evaporated to obtain the
total alkaloidal fraction. Moreover, total alkaloidal fraction
was able to further liquid-liquid partition to aord weak

Evidence-Based Complementary and Alternative Medicine 
HO
O
HO
O
H
O
O
O
N
O
O
O
N
O
12
6
12
8
N
H
O
O
OH
O
N
OH
14 15
N
O
OH
N
H
O
O
O
13
16
OH HO O
OO
N
17
22
10 H
11 AC
R
O
OH
R
O
O
N
18
19 OH
R
O
OH
OH
O
O
20
21
N
O
HO
O
O
9
7
N
OH
O
O
OH
H3
N
H
N
N
H
H
45
N
O
N
N
O
NH
N
N
H
O
N
N
H
O
N
O
N
HOR N
N
H
O
R1
R1
R2
R2
R3
R3
OCH3
CH3CH3CH3
CH3CH3CH2OH
N+
N+
F : Chemical structures of alkaloids (–)fromgenusCaulophyllum.
base (Fr. ), nonphenolic alkaloids (Fr. ), and phenolic
alkaloids (Fr. ) []. e H2O layer was neutralized with
% HCl and extracted with n-butanol. e combined organic
layers were evaporated to obtain total saponin fraction [].
Column chromatography is also a popular method to enrich
total alkaloids and saponins from Caulophyllum plants by
choosing optimal macroporous or (and) ion exchange resins
[,,].
2.1. Alkaloids. With respect to alkaloid aspects of this genus,
 molecules have been isolated and identied from genus
Caulophyllum. Alkaloid compounds are very important bio-
active constituents in genus Caulophyllum.eirchemical
structuresandsourcescanbeseeninFigureand Table .
ese compounds can be divided into several kinds of struc-
tural types. magnoorine (), taspine (), and boldine ()are
contributed to aporphine alkaloids. Aporphine alkaloids have
beenshowntopossessanticanceractivityandthereisevi-
dence that this activity is exerted through induction of apop-
tosis, inhibiting cell proliferation and inhibiting DNA topoi-
somerase [,]. Magnoorine (), a quaternary ammonium
base, is isolated and detected with the biggest amounts among
all the alkaloids isolated from genus Caulophyllum.was also
isolated from the n-butanol fraction of blue cohosh due to its
strong water-solubility, but it was not active in the rat embryo
culture []. e molecular structure of is characterized by
high symmetry. – are typical quinolizidine alkaloids.

 Evidence-Based Complementary and Alternative Medicine
T : Chemical structures of alkaloids (–)fromgenusCaulophyllum.
No. Compounds Formula SourcesaReferences
Magnoorine CH NO+Cr, Ct [,]
Taspine CH NOCr, Ct [,]
Boldine CH NOCr []
Anagyrine C HNOCr, Ct [,,]
Sparteine CH NCr, Ct []
N-methylcytisine CH NOCr, Ct [,–]
Cytisine C H NOCr []
,-Dehydro-𝛼-isolupanine C H NOCr, Ct [,]
Lupanine CHNOCr, Ct [,]
 Baptifoline or Argentamin CH NOCt [,]
 O-acetylbaptifolin CHNOCt []
 𝛼-isolupanine CH NOCr, Ct [,]
 alictroidine CH NOCt [,]
 Caulophyllumine A CH NOCt []
 Caulophyllumine B CH NO Ct []
 Piperidylacetophenone CH NOCt []
 Caulophine CH NOCr []
 Caulophylline A CH NOCr []
 Caulophylline B CH NOCr []
 Caulophylline C CH NOCr []
 Caulophylline D CHNOCr []
 Caulophylline E CH NOCr []
aCr means C. robustum;Ct means C. thalictroides.
Quinolizidine alkaloids have been reported to possess the
obvious nematicidal activity [].
In October , a novel alkaloid, thalictroidine ()with
piperidine-acetophenone conjugate, was isolated from the
rhizomes of C. thalictroides using an in vitro rat embryo cul-
ture method.  was not teratogenic in the rat embryo culture
at tested concentrations []. Aer nine years,  was isolated
again from C. thalictroides,togetherwith– []. –,
piperidine-acetophenone conjugates, are rare in the plant
kingdom.  was only reported from Boehmeria genus []
and is another example of such a type of compound from
natural sources.
In April , a distinct class of alkaloid, uorenone
alkaloid (caulophine, ), was rstly reported from the radix
of C. robustum using cell membrane chromatography as the
screening method.  was identied as -(-(dimethylamino)
ethyl)-,-dihydroxy-,-dimethoxy-H-fluoren--one based
on physicochemical and spectroscopic analyses.  pos-
sessed antimyocardial ischemia activity by rat experiments.
It is worth mentioning that a preparative high performance
liquid chromatography method was developed for isolation,
purication, and enrichment of caulophine ()[]. As
follows, another four uorenone alkaloids, caulophyllines A–
D(–), and one dihydroazauoranthene alkaloid, caulo-
phylline E (), were isolated from the roots of C. robustum.
Fluorenone alkaloid is a newly discovered alkaloid skele-
toninnaturalproducts.–, ve new uorenone alkaloids,
were isolated from the same plant, suggesting that uorenone
type alkaloid is another kind of metabolites that existed in this
genus Caulophyllum. is a novel and rare naturally occurred
dihydroazauoranthene alkaloid, there are no reports about
dihydroazauoranthene alkaloid isolated from natural prod-
ucts except its novel core skeleton rst isolated from coal tar
[].  has the isoquinoline fragment, which is possible to be
the conceivable precursor of dierent substituted uorenone
alkaloids. A hypothetical biosynthetic pathway for  was
proposed starting from , which undergoes a sequential
nitrogen-related double bond reduction, oxidation, ring-
opening, N-methylation, and demethoxy process [].
2.2. Triterpene Saponins. Caulophyllum triterpenes generally
constitute the main class of secondary metabolites in the
genus Caulophyllum amountingtoupto.%ofthetotaldry
weight in root and rhizome []. Until now,  caulophy-
llsaponinswereisolatedandidentiedbychemicaland
detailed spectroscopic analysis (Table ). ese saponins
generally bear one (monodesmosidic) or two (bidesmosidic)
carbohydrate chains that are directly attached to the hydroxyl
groups in position C- for monodesmosidic saponins and to
positions C- and C- in the case of the bidesmosidic
saponins. ,,,,, –, and – are bidesmo-
sidic triterpenoid saponins with two sugar chains at C- and
C-. Others were found to only have one sugar chain at C-
or C-.
Twelve kinds of aglycones have been discovered from the
genus Caulophyllum (Figure (a))[,,]. Before ,
only four kinds of sapogenins were discovered from genus

Evidence-Based Complementary and Alternative Medicine 
T : Chemical structures of triterpene saponins (–)fromgenusCaulophyllum.
No. Compound names C- C- Formula SourcesaReferences
 Ara →𝛽-O-AGS—C
H OCt []
 Saponin PE, Glc →Ara →𝛽-O-AGS—C
H O Ct [,]
 Ciwujianoside A, Glc →Ara →𝛽-O-AG --O←Glc←Glc←Rha SSC HO Ct [,]
 Cauloside A, Ara →𝛽-O-AGS—C
H OCr, Ct [,,,,
,]
 Cauloside C, Glc →Ara →𝛽-O-AGS—C
H O Cr, Ct [,,,,
]
 Cauloside D, Ara →𝛽-O-AG--O←Glc←Glc←Rha SSCH O Cr, Ct [–
,,]
 Cauloside G, Glc →Ara →𝛽-O-AG--O←Glc←Glc←Rha SSC H O Cr, Ct [–
,,]
 Cauloside b, Rha →Ara →𝛽-O-AG S—C
H O Cr []
 Cauloside c, Ara →Rha →Ara →𝛽-O-AGS—C
H O Cr []
 AG--O←Glc←Glc←Rha —S
CH O Ct []
 Glc →Ara →𝛽-O-AG--O←Glc←Glc←Rha SSCH O Cr [,]
 Ara →𝛽-O-AG--O←Glc←Glc←Rha SSCH O Ct []
 Ara →𝛽-O-AGS—C
H OCt, Cr [,,]
 Glc →Ara →𝛽-O-AGS—C
H O Ct [,]
 Leiyemudanoside C, Glc →Ara →𝛽-O-AG --O←Glc←Glc←Rha SSCH O Cr []
 Ara →𝛽-O-AG--O←Glc←Glc SSCH O Ct []
 Glc →Ara →𝛽-O-AG--O←Glc←Glc←Rha SSCH O Cr []
 Cauloside B, Ara →𝛽-O-AG S—C
H OCr, Ct [–,,
,,]
 Leonticin D, Ara →𝛽-O-AG --O←Glc←Glc←Rha SSC HO Ct [,,]
 Cauloside H, Glc →Ara →𝛽-O-AG--O←Glc←Glc←Rha SSC H O Ct []
 Leiyemudanoside A, Ara →𝛽-O-AG --O←Glc←Glc SSC H O Cr []
 Leiyemudanoside B, Glc →Ara →𝛽-O-AG--O←Glc←Glc←Rha SSCH O Cr []
 Glc →Ara →𝛽-O-AGS—C
H O Ct []
 Ara →𝛽-O-AG--O←Glc SSCH O Ct []
 Ara →𝛽-O-AGS—C
H OCt []
 Ara →𝛽-O-AGS—C
H OCt []
 Glc →Ara →𝛽-O-AGS—C
H O Ct []
 Ara →𝛽-O-AGS—C
H O Ct []
 Ara →𝛽-O-AGS—C
H OCt []
 Glc →Ara →𝛽-O-AG S—C
H O Ct []
 Glc →Ara →𝛽-O-AG S—C
H O Ct []
 Ara →𝛽-O-AG S—C
H OCr []
aCr means C. robustum;Ct means C. thalictroides.
Caulophyllum,namely,oleanolicacid(AG
1), hederagenin
(AG2), echinocystic acid (AG3), and caulophyllogenin (AG4).
However, Ma et al. reported – with abnormal sapogenins
AG5to AG11 from blue cohosh for the rst time. As follows,
 bearing sapogenin erythrodiol was discovered from genus
Caulophyllum in  []. ese aglycones are closely related
oxygenated pentacyclic triterpenoidal structures that can be
distinguished only by the positions and numbers of the dou-
ble bonds in rings C and D and oxygenation patterns in posi-
tions C-, C-, and C-. A possible biosynthetic pathway
of Caulophyllum sapogeninscanbehypothesized,asshownin
Scheme .erstis,that,-oxidosqualeneiscyclizedtothe
pentacyclic oleanane-type triterpenoid backbone 𝛽-amyrin
by plant oxidosqualene cyclases 𝛽-amyrinsynthase[,].

 Evidence-Based Complementary and Alternative Medicine
HO
COOH
HO
COOH
OH
HO
COOH
OH
HO
COOH
OH
OH CHO
HO
COOH
OH
CHO
HO
COOH
HO
OH
OOO
HO
OH
OH
OOO
HO
COOH
OH
OH
HO
COOH
H
OH
HO
H
O
O
H
HO
OH
OH
AB
CD
E
1
3
12
16 28
23
O
AG1AG2AG3
AG4AG5AG6
AG7AG8AG9
AG10 AG11 AG12
CH2OH
(a)
O
O
O
O
O
O
OH
O
O
O
O
O
HO
HO
HO
HO
HO
HO
HO
HO
HO
HO
HO
HO
HO
HO
HO
HO
HO HO
HO HO
HO
HO
HO
HO
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
O
O
O
O
O
O
O
O
O
O
O
OO
S1S2S3S4
S5S6S7S8
(b)
F : (a) Chemical structures of aglycones (AG) of saponins; (b) linkage mode of sugar moieties of saponins from genus Caulophyllum.
e 𝛽-amyrin experienced hydroxylation at C- to produce
erythrodiol (AG12). Erythrodiol is further oxidized at the C-
 position by a single cytochrome P enzyme to yield
oleanolic acid (AG1)[]. e aglycones AG1–AG9may be
derived from the common skeleton of oleanolic acid as
precursors that rstly experience selective oxidation at C-,
or C-, or both of C- and C- to aord hederagenin
(AG2), echinocystic acid (AG3), and caulophyllogenin (AG4),
respectively. Hederagenin may selectively involve a complex
process such as dehydrogenization, oxidation, lactonization,

Evidence-Based Complementary and Alternative Medicine 
O
2,3-oxidosqualene
HO HO
OH
HO
OH
O
HO
OH
O
OH
HO
OH
O
OH
HO
OH
O
OH
OH
HO
OH
O
H
O
HO
OH
O
OH
H
O
HO
OH
O
OH
HO
OH
OOO
HO
OH
OOO
HO
OH
O
OH
OH
OH
OHOH
HO
OH
OO
OH
HO
OH
OO
AG1
AG2
AG3AG4
AG5AG6
AG7
AG8
AG9
AG12
𝛽-Amyrin
S : Hypothesized biosynthetic pathway for oleanane aglycones from genus Caulophyllum. A series of step-by-step actions from ,-
oxidosqualene to 𝛽-amyrin, erythrodiol, oleanolic acid, and other aglycones are assumed.
dehydration, and lactone ring hydrolysis to form diverse
aglycones in genus Caulophyllum. ough the intermediate
(𝛽,𝛼-dihydroxy-olean--oic acid 𝛾-lactone) has been
articially synthesized from oleanolic acid (Supplemen-
tary Scheme  in Supplementary Material available online
at http://dx.doi.org/.//)[], this type of
biosynthetic pathway in plants also needs to be further
conrmed.
e according carbohydrate chains are composed mainly
of arabinose, rhamnose, and glucose moieties. Aer acid
hydrolysis, gas chromatography analysis revealed the pres-
ence of glucose, arabinose, and rhamnose through comparing

 Evidence-Based Complementary and Alternative Medicine
OH
O
55
RO
R
56 H
57 Glc
HO 59
HO
58
HO
60
F : Chemical structures of other compounds (–)fromgenusCaulophyllum.
with derivatives obtained by the same method of standard
monosaccharides [].Fortheirlinkagemodeofsugar
moieties, it showed the presence of many linked types of sugar
moieties, including terminal glucose, ,-linked glucose, ter-
minal rhamnose, ,-linked rhamnose, terminal arabinose,
,-linked arabinose, ,-linked arabinose, and ,,-linked
arabinose [,,,](Figure(b)). Diverse aglycones,
monosaccharide residues, and diverse linkage mode of sugar
moieties are possible to form diverse structures of triterpene
saponins from genus Caulophyllum.
2.3. Other Compounds. Other minor compounds are present
in this genus, such as fatty acids and sterols [,]. ese
compounds were identied as palmitic acid (), 𝛼-
spinasterol (), 𝛼-spinasterol-𝛽-D-glucopyranoside (),
stigmasterol (), lupeol (), and cholesterol ()(Figure).
Polysaccharides are present in this genus. e extraction
process and antioxidant activity of polysaccharides have been
studied [,]. e optimal ultrasound-assisted extraction
conditions of polysaccharides from C. robusutm were extr-
action temperature ∘C, time  min, ratio of liquid to solid
, and extracting power of  W.
3. Synthetic Taspine Derivatives
Zhang et al. designed and synthesized four novel ring-opened
target compounds (1󸀠–4󸀠) by structure-based drug design.
is design includes two pathways: cleavage of the C–C
bond of diphenyl and ester bond of ring B and ring D
(Scheme (A)). Targeted compounds 1󸀠and 2󸀠were synthe-
sized by the route outlined in Scheme (B). Isovanillin (5󸀠)
was used as the starting material, which was rstly oxidized
to aord isovanillic acid (6󸀠). e methyl ester 7󸀠was pre-
pared to avoid side-reactions of the carboxylate group. en
reuxing of 7󸀠with prenylbromide in the presence of K2CO3
in anhydrous acetone aorded 8󸀠. A solution of 8󸀠in N,N-
dimethylaniline was heated to reux to give 9󸀠. Prenyl group
was moved into the paraposition of hydroxyl in Claisen rear-
rangement process. e next step was the coupling of 9󸀠to the
carboxyl of 10󸀠by an ester bond with DCC and DMAP. e
oxidation of 11󸀠produced aldehyde 12󸀠, which reacted with
dimethylaminefollowedbyreductiontogive13󸀠.Atlast,ben-
zyl deprotection of 13󸀠with palladium-carbon in MeOH gave
1󸀠.e3󸀠and 4󸀠were synthesized in the same way from iso-
vanillin [].
Synthetic endeavors into cleavage of the C–C bond
andesterbondofringsB,D,andEhavebeenstudied
(Scheme (A)). Initially, six target biphenyl derivatives (19󸀠–
24󸀠) were successfully synthesized by general routes des-
cribed in Scheme (B) employing a classical symmetrical
Ullmann reaction []. Isovanillin (5󸀠)wasalsousedforthe
starting material, which was required for seven steps to aord
19󸀠by bromination, benzylation, oxidation, substitution
reaction, Ullmann reaction, and catalytic hydrogenation. 19󸀠
is an important intermediate to synthesize the following tar-
geted compounds. During the synthesis of unsymmetrical
biphenyl (22󸀠), a novel symmetrical biphenyl derivative (31󸀠)
was surprisingly isolated as a byproduct [], which exhibited
potent anticancer activity to attract increasing attention. To
further investigate this nding, researchers aimed to enhance
the structural complexity and diversity of 22󸀠by generating
novel biphenyls (Figure )[]. As a result, eighteen sym-
metrical biphenyls derivatives (31󸀠–48󸀠) were rstly prepared
[]. Following these, He et al. used 20󸀠as the identifying
group and synthesized another two novel taspine diphenyl
derivatives 49󸀠and 50󸀠, which were made by introducing
coumarin groups into the structure of 20󸀠[]. Meanwhile,
derivatives 51󸀠and 52󸀠were obtained via similar procedures
(Scheme (B)) [].

Evidence-Based Complementary and Alternative Medicine 
O
O
O
N
O
O
O
A
BD
C
Taspine 2
COO
OH
O
R
O
O
O
O
O
HO
OOC
N
O
O
OH
O
OH
O
ab
OH
O
c
O
O
COO
d
OH
O
COO
+
OBn
O
e
COO
OBn
O
O
O
O
f
COO
OBn
O
O
OO
g
COO
OBn
O
O
O
ON
h
i
BnO
O
COO
j
BnO
O
OH
O
+e
O
O
HO
OOC
O
O
fO
O
HO
OOC
O
O
O
O
O
BnO
OOC
N
O
O
gh
(A)
(B)
D
B
O
H
O
OH
O
O
O
H
O
OH
O
O
1
󳰀
1
󳰀
R=N(CH3)2
3󳰀R=morpholine
4󳰀R=3-Cl-4-F-aniline
5󳰀6󳰀7󳰀
7󳰀
8󳰀9󳰀
9󳰀
9󳰀
10󳰀
11
󳰀12󳰀13󳰀
14󳰀
15󳰀16󳰀
16󳰀
17󳰀18󳰀
OCH3
2󳰀
2󳰀
S : (A) Design of ring-opened target compounds 1󸀠–4󸀠;(B)preparationoftargetcompounds1󸀠–4󸀠. Reagents and conditions: (a)
NaOH, KOH, H2O, %; (b) CH3OH, H2SO4, %; (c) prenyl bromide, K2CO3, acetone, %; (d) N, N-dimethylaniline, N2,reux,%;(e)
anhydrous THF, DCC, DMAP, %; (f) OsO4,NaIO
4, acetone/H2O/t-BuOH, %; (g) dimethylamine /Morpholine/-Cl--F-aniline, THF,
CH2Cl2, NaBH(OAc)3, –%; (h) H2, Pd/C, %; (i) BnCl, K2CO3, EtOH, %; (j) NaOH/H2O, CH3OH, % [].
4. Bioactivity
4.1. Antibacterial Activity. Earlier biological studies showed
that caulosides A–D and G (,,–) have antimicrobial
activity [].Recently,triterpenecompoundsisolatedfromC.
robustum showed microorganism inhibitory activities to the
test fungi and bacteria. Moreover, compound  and caulo-
side B () had notable inhibiting microorganism activities
to bacteria with minimal inhibitory concentration (MIC) of
. 𝜇g/mL []. Ethanol extract and its ve subfractions of C.
robustum showed high antibacterial activity against Staphy-
lococcus aureus,Staphylococcus aureus (clinic bacterial), and

 Evidence-Based Complementary and Alternative Medicine
O
O
O
N
O
O
O
Taspine 2
CONH
HNOC
O
O
(A)
E
A
B
C
DB, D, E
H
N
OF
Cl
N
O
N
HN
N
O
OH
O
H
a
O
OH
O
Hb
Br O
OBn
O
Hc
Br O
OBn
O
OH d
Br O
OBn
O
Cl e
Br
O
OBn
O
NH f
Br
O
O
OBn
CONH
BnO
HNOC
gh
(B)
or
or
or
By products
i
j
R1=H, R2=H
R1=W, R 2=H
R1=H, R2=Y
R1=W, R 2=Y
R1=W, R 2=X
R1=W, R 2=Z
OR2
R1O
W=
X=
Y=
Z=
51
󳰀
52󳰀49󳰀+50󳰀
32󳰀–48󳰀
31
󳰀
20󳰀
20󳰀
30󳰀
29󳰀
19
󳰀
23󳰀
19
󳰀
21
󳰀
21
󳰀
22󳰀
22󳰀
23󳰀
24󳰀
24󳰀
25󳰀26󳰀27󳰀28󳰀
5󳰀
S : (A) Design of ring-opened target compounds 19󸀠–24󸀠; (B) preparation of target compounds (19󸀠–24󸀠). Reagents and Conditions:
(a) Fe, NaOAc, AcOH, Br2,%;(b)BnCl,K
2CO3,%;(c)NaH
2PO4,NaClO
2,%H
2O2,%;(d)SOCl
2, DMF(cat), CH2Cl2, %; (e)
CH2Cl2,%CH
3NH2,%;(f)Cu,DMF,%;(g)H
2, Pd/C; %; (h) K2CO3,DMF,(,%;,%);(i)K
2CO3,EtOH,(,%;,
%; , %) [,–].
Bacillus subtilis, and the diameters of the biggest inhibition
zone were . mm, . mm, and . mm. e MICs of
these were .–.mg/mL [].
4.2. Anti-Inammatory and Analgesic Eects. e anti-
inammatory and analgesic eects of ethanol extract, chlo-
roform extract, and n-butyl alcohol extract from C. robustum
were observed by several animal experiments. Among the dif-
ferent organic extracts, the action of alcohol extract was better
than other organic extracts []. Cauloside A ()andcaulo-
side C () had anti-inammatory and analgesic activities at
dose dependency and the analgesic eect was the most signif-
icant when compounds were injected for  min []. From
the points of structure-activity relationship of the saponins,
cauloside C () with disaccharide has more potent analgesic
eect than cauloside A () with monosaccharide. Oppo-
sitely, cauloside A () has more potent anti-inammatory
activity than cauloside C (). e anti-inammatory activity
of taspine hydrochloride has been demonstrated by using the
carrageenan-induced pedal edema method, the cotton pellet-
induced granuloma method, and the adjuvant polyarthritis
model [].
Lee et al. () assessed the in vitro and in vivo eects of
blue cohosh on lipopolysaccharide (LPS)-induced cytokines
in BV cells and mice. Several lines of evidence indicate that
blue cohosh treatment suppressed the elevation of LPS-
induced iNOS (inducible nitric oxide synthase) expression in
a concentration-dependent manner in microglia cells. Blue
cohosh saponins (caulosides A−D: ,,,)signi-
cantly suppressed the expression of tumor necrosis factor-𝛼
(TNF-𝛼), interleukin-𝛽(IL-𝛽), and IL-. In addition, blue
cohosh extract suppressed the expression of COX (cyclo-
oxygenase)-, iNOS, and proinammatory cytokines in
adrenalglandsofmice.So,itisconcludedthatsaponin
constituents of blue cohosh exert anti-inammatory eects
through the inhibition of expression of iNOS and proinam-
matory cytokines [].
4.3. Antioxidant Eects. Caulophylline A−D(–) aorded
the lower scavenging eects against DPPH (,-diphenyl--
picrylhydrazyl) radical at test concentration (. to .
𝜇g/mL). Caulophylline E () showed good scavenging ee-
cts against DPPH radical with IC50 (half-inhibition con-
centration) of . 𝜇g/mL []. Antioxidant activities of the

Evidence-Based Complementary and Alternative Medicine 
O
O
O
O
H
N
O
C
N
H
O
HN
NH
a
b
c
d
e
f
g
h
i
j
Isop a
Isop b
Isop c
Isop d
Isop e
Isop j
Diet a
Diet b
F
Cl
Cl
Cl
O
Br
F
NF
F
O
O
O
HNOC
O
O
OOO
NH
O
F
Cl
O
O
HNOC
CONH
CONH
O
O
OO H
N
NH
O
F
Cl
O
O
N
H
O
NH2
NH
O
O
F
Cl O
O
H
N
O
F
Cl
O
H
N
O
OH
O
NH
O
O
NH
NH
O
O
31
󳰀
32󳰀
33󳰀
34󳰀
35󳰀
36󳰀
37󳰀
38󳰀
39󳰀
40󳰀
41
󳰀
42󳰀
43󳰀
44󳰀
45󳰀
46󳰀
47󳰀
48󳰀
a=
b=
c=
d=
e=
CF3
f=
g=
h=
i=
j=
49󳰀
50󳰀
51
󳰀
52󳰀
R1
R1
R1
R2
R2
R2
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
Isop =Isopropyl; Diet =Diethyl
F : Structures of 31󸀠–51󸀠.
polysaccharide fraction, ethanol extract, and dierent polar
fractions of C.robustum were evaluated by DPPH, hydroxyl,
and superoxide radical and nitrogen dioxide (NO2) scaveng-
ing assay [,]. e results showed that ethanol extract and
dierent polar fractions displayed high antioxidant activities.
e scavenging activities of polysaccharides from C. robus-
tum for DPPH, hydroxyl, and superoxide radical and NO2
were attributed to %, %, %, and .%, respectively,
for the concentrations of . mg/mL. Another experiment
research reported that chloroform partition fraction showed
IC50 value of DPPH-free radical-scavenging activity which
was . 𝜇g/mL [].
4.4. Antiacetylcholinesterase Activity. As early as , tasp-
ine () has been conrmed to be an antiacetylcholinesterase
(AChE) inhibitory agent by a bioactivity-guided approach
in a Magnolia x soulangiana extract using a microplate
enzyme assay with Ellman’s reagent []. showed a sig-
nicantly higher eect on AChE than the positive control
galantamine and selectively inhibited the enzyme in a long-
lasting and concentration-dependent fashion with an IC50
value of . 𝜇g/mL. It could be suggested that taspine might
be a potential candidate for the development of anti-AD
(Alzheimer’s disease) treatment.
More recently, C. robustum has been conrmed to possess
signicant AChE activity with inhibition rates (. ±.)%
at the concentration of  g⋅L−1 through thin layer chromatog-
raphy bioautographic method. Furthermore, chloroform
fractions have shown higher AChE inhibitory capacity, so
it will be further performed bioguided isolation and purica-
tion to obtain active compounds []. In addition to taspine in
C.robustum, whether to have other compounds responsible
for the activity of AChE is worthy of studying further.
4.5. Eect on Atherosclerosis and Myocardial Ischemia. It was
found that the n-butanol fraction of C.robustum was an eec-
tive part, and caulophine () separated from the part was an
activeoneinvasodilatation[,]. e n-butanol fraction
may have protective action on H2O2injured-human umbil-
ical vein endothelial cell line in vitro, and its mechanism of
actionmayberelatedtotheincreaseofthelevelofnitricoxide
(NO), NOS (nitric oxide synthase), and the expression of
NF-𝜅B (nuclear factor kappa B) []. e interaction between

 Evidence-Based Complementary and Alternative Medicine
the eective component  and the membrane or membrane
receptor was reected in the vascular CMC model, which
suggested that  may exert bioactivity in the heart [].
e deeper study demonstrates that  is able to protect
cardiomyocytes from oxidative and ischemic injury through
an antioxidative mechanism [] and from caeine-induced
injury via calcium antagonism [].
4.6. Antitumor Activity and Mechanism of Action. e cyto-
toxicity (IC50)oftaspine()wasfoundtobe.𝜇g/mL
against KB cells and . 𝜇g/mL against V- cells []. 
showed antitumor activity on the mouse S sarcoma in a
good dose-dependent manner []. e inhibition rates on
tumor of taspine at low, middle, and high concentrations were
.%, .%, and .%, respectively. e microvessel
density and protein expressing of the vascular endothelial
growth factor (VEGF), basic broblast growth factor (bFGF),
Bcl-, and Bax in the tumor were decreased compared with
thenegativecontrol.eratioofBaxtoBcl-wasincreased.
has antitumor eect on the S sarcoma, and the mechanism
may be through the way of decreasing the expressing of
theVEGF,bFGF,Bcl-,andBaxandinducingthevascular
endothelial cell apoptosis.
Zhan et al. was to investigate the eect of taspine on
the growth of oestrogen-receptor-positive breast cancer
xenogras in vivo and the possible mechanism for this
action []. Cell cycle and apoptosis analysis documented that
taspine was able to change cell cycle and induce cell apoptosis.
ere was a signicant decrease in the expression of estrogen
receptor (ER) and progesterone receptor (PR) both in tumor
tissue and cells aer treatment with taspine. At the same time,
it also showed a reduction in the expression of mRNA for
ER and PR in the group treated with taspine. ese data
suggested that taspine might serve as a promising candidate
of ER antagonist in the treatment of oestrogen-independent
breast cancer.
Many evidences have shown that taspine could suppress
tumor-induced angiogenesis. Taspine was able to inhibit
chicken chorioallantoic membrane angiogenesis through
interfering with the proliferation and migration of endothe-
lial cells in a dose-dependent manner []. e exact mech-
anism [] has been further demonstrated, suggesting that
VEGF and bFGF secretion were downregulated by taspine in
human non-small cell lung cancer cell (A cell) and human
umbilical vein endothelial cells (HUVECs), conrmed by
the decreased mRNA level of VEGF and Flk-/KDR aer
taspine treatment in HUVECs. e molecular mechanisms
of taspine on tumor angiogenic inhibition have been further
studied in vitro [], which indicated that taspine signicantly
inhibited cell proliferation of HUVECs induced by VEGF
via decreasing Akt and Erk/ activities except decreasing
VEGF level. Authors assume that taspine can inhibit the pro-
liferation of vascular endothelial cells in tumor by regulating
PI kinase and MAP kinase signal pathways.
Additionally, taspine could induce apoptosis of HUVECs
in a dose-dependent manner []. Cell cycle was signi-
cantlystoppedattheSphase.emorphologyofHUVEC
treated with taspine showed nuclear karyopycnosis, chro-
matin agglutination, and typical apoptotic body detected by
electronic microscope. Taspine has an inhibitory eect on
growth of HUVECs and can induce its apoptosis by decreas-
ing Bcl- expression and increasing bax expression. Zhang
et al. continuously investigated the eects of taspine on the
proliferation and apoptosis in the A cell []. e cell cycle
was signicantly stopped at S phase, and nuclear karyopy-
knosis, chromatin agglutination, and typical apoptotic bodies
were found aer taspine treatment in A cells. ere
was a decrease in the expression of Bcl-, whereas the expres-
sion of caspase-, cleaved caspase-, CDK, CDK, and Bax
increased. ese data demonstrated that taspine can induce
apoptosis by activating caspase- expression and upregulat-
ing the ratio of Bax/Bcl- in A cells.
e preliminary biological test demonstrated that deriva-
tive 3󸀠showed much better inhibitory activities against
CACO- (IC50 =.𝜇g/mL) and ECV (IC50 =
. 𝜇g/mL) than taspine []. A deep research demon-
strated that most derivatives (1󸀠–4󸀠and 19󸀠–24󸀠)possesseda
moderate degree of cytotoxicity against human cancer cell
lines []. One of them (3󸀠) exhibited much better antipro-
liferative activity against CACO- (IC50 = . 𝜇g/mL) and
ECV (IC50 =.𝜇g/mL) cells than taspine did. Some
of the compounds showed good antiproliferative activity
against colon (HT), breast (MCF-), lung (A), rectum
(CACO-), skin (A), hepatoma (), and pancreatic
(PANC-) cancers cell lines. A continual research demon-
strated that derivative 3󸀠can inhibit the proliferation of, and
induce apoptosis in, Caco- cells by activating caspase-,
caspase-, and caspase-, downregulating the expressions of
VEGF, and upregulating the ratio of bax/bcl- [].
Derivatives (31󸀠)and(32󸀠) demonstrated the most potent
cytotoxic activity with IC50 values between . 𝜇g/mL and
. 𝜇g/mL among symmetrical taspine derivatives (31󸀠–48󸀠)
[]. Biphenyls without halogen substitution (34󸀠,38󸀠,39󸀠,
and 44󸀠) were much less potent than those containing
halogen. Halogen substitution played a critical role in the
activity of biphenyls. Derivative 31󸀠inhibits tumor growth in
xenograedAcellsinnudemicebyinhibitingthegrowth
of neovessels. In other words, derivative 31󸀠is an inhibitor of
angiogenesis which functions by downregulating VEGF [].
Furthermore, derivative 31󸀠had potential to suppress the
adhesion, migration, and invasion of ZR-- cancer cells,
and it could serve as a potential novel therapeutic candidate
for the treatment of metastatic breast cancer []. Derivative
48󸀠could inhibit proliferation of lovo cell and tumor growth
in a human colon tumor xenograed model of athymic mice,
which might be a novel angiogenesis inhibitor that reduces
angiogenic responses in vivo and in vitro by blocking VEGFR
signaling pathways [].
Two novel derivatives (49󸀠and 50󸀠)byintroducingdier-
ent coumarin uorescent groups into the basic structure have
notonlyuorescencebutalsotheabilitytoinhibiteectson
dierent breast cancer cell lines, which indicates their pos-
sible further use as dual functional uorescence probes in
tracer analysis. Derivative 51󸀠inhibits tumor growth and cell
proliferation by inhibiting cell migration, downregulating
mRNA expression of VEGF and EGF, and decreasing angio-
genic factor production, which deserves further consid-
eration as a chemotherapeutic agent []. All evidences

Evidence-Based Complementary and Alternative Medicine 
have demonstrated that the lactone ring B is important for
activity, while the lactone ring D can be opened, thus retain-
ing and even improving the antiproliferative properties of
taspine. Halogen substitution could potentially improve the
anticancer activity of the biphenyl derivatives [,,].
4.7. Inhibitory Cytochrome P450 Eects. e methanolic extr-
acts of the roots of blue cohosh, the alkaloidal fraction,
andisolatedconstituentswereevaluatedfortheirinhibition
of major drug metabolizing cytochrome P (CYP)
enzymes []. e methanolic extracts did not show any
eect but the alkaloidal fraction showed a strong inhibition of
CYPC, A, D, and A (>% inhibition at  𝜇g/mL)
with IC50 values in the range of – 𝜇g/mL. Among the
isolated alkaloids, caulophyllumine B (), O-acetlybaptifolin
(), anagyrine (), and lupanine () inhibited these enzymes
to various extents (IC50: .–. 𝜇g/mL). N-methylcytisine
() showed weak activity against the CYPA in vitro with
% inhibition at . 𝜇g/mL. An equimolar mixture of
alkaloids exhibited a more pronounced inhibitory eect on all
fourenzymesascomparedtotheisolatedalkaloids.Among
the saponins, caulosides C ()andD()showed%and
%inhibitionofCYPAattheconcentrationof.and
. 𝜇g/mL, respectively. Other enzymes were not aected.
is in vitro study indicates that dietary supplements contain-
ing blue cohosh may pose a risk of drug-drug interactions if
taken with other drugs or herbs, metabolism of which
involves CYP enzymes.
4.8. Topoisomerase Inhibitor. Taspi n e ( ) was found to induce
conformational activation of the proapoptotic proteins Bak
and Bax, mitochondrial cytochrome c release, and mitochon-
drial membrane permeabilization in HCT cells []. Anal-
ysis of the gene expression signature of taspine treated cells
suggested that taspine is a topoisomerase inhibitor. Taspine
has a reduced cytotoxic eect on a cell line with a mutated
topoisomerase II enzyme. Interestingly, in contrast to the top-
oisomerase II inhibitors doxorubicin, etoposide, and mitox-
antrone, taspine was cytotoxic to cell lines overexpressing the
PgP or MRP drug eux transporters. Taspine induces wide-
spread apoptosis in colon carcinoma multicellular spheroids
and that apoptosis is induced in two xenogra mouse models
in vivo. Taspine is a dual topoisomerase inhibitor that is
eective in cells overexpressing drug eux transporters and
induces wide-spread apoptosis in multicellular spheroids.
4.9. Eect on Wound Healing. A patent reported that the
method is useful for preparing wound care composition,
which comprises C. robustum,whichisusefulforrelieving
postoperative pain and promoting wound healing and blood
circulation in wound area []. Further research showed that
taspine was able to promote early phases of wound healing in
a dose-dependent manner with no substantial modication
thereaer. Its mechanism of action is probably related to its
chemotactic properties on broblasts and is not mediated
by changes in extracellular matrix []. Authors summarized
that taspine opens a pathway of research for new tools to stim-
ulate wound repair in the absence of macrophages, thereby
helping to better understand the process of wound healing.
Taspine also exhibited a dose-related cicatrizant eect and a
median eective dose (ED50) of . mg/kg, which was
nontoxic to human foreskin broblasts at concentrations
below  ng/mL and that had no eect on cell proliferation
[].
4.10. Toxicity. N-methylcytisine () exhibited teratogenic
activity in the rat embryo culture (REC), an in vitro method to
detect potential teratogens. Anagyrine ()and𝛼-isolupanine
() were not teratogenic in the REC at tested concentrations.
Taspin e ( ) showed high embryotoxicity, but no teratogenic
activity, in the REC []. Wu et al. have observed that blue
cohosh interrupted medaka embryogenesis and produced an
abnormal phenotype, which identies blue cohosh as a potent
teratogen. Moreover, the induction of gata mRNA followed
by edn mRNA by BC indicates that the teratogenic response
of blue cohosh is probably mediated by the Gata-End
signaling pathway []. Caulosides B ()andC()were
reported to have cytotoxicity to developing sea urchin
embryos by changing cell permeability. It is well-known that
cytotoxic glycoside causes a disturbance of cell membrane
permeability that can cause leakage of important cellular
components [,].
A new born infant whose mother ingested an herbal med-
ication, blue cohosh, to promote uterine contractions pre-
sented with acute myocardial infarction associated with pro-
foundcongestiveheartfailureandshock[]. One year later,
other similar cases were reported []. Meanwhile, According
to a survey of midwives in the United States, approximately
% of midwives reported using blue cohosh as a labour-
inducing aid. Severe multiorgan hypoxic injury may occur.
Recently, a review focused on the toxicity of blue cohosh has
been reported [].
5. Pharmacokinetics
Magnoorine (), taspine (), and caulophine ()werethe
main components of genus Caulophyllum.Severalstudies
have been carried out to understand the distribution, absorp-
tion, metabolism, and excretion of magnoorine (), taspine
(), and caulophine () using modern analytical methods.
5.1. Pharmacokinetics of Magnoorine. As far as magnoo-
rine is concerned, a new sample-preparation method based
on hollow-ber liquid-phase microextraction (HFLPME)
was developed and successfully used for pharmacoki-
netic studies of magnoorine in rat plasma aer intra-
venous administration. e magnoorine disappears from
rat plasma in accordance with a two-compartment open
model. e plasma concentration of magnoorine reached a
peak immediately aer completion of administration, then
began to decline. Without doubt, the chromatographic and
HFLPME sample-preparation procedures of magnoorine
will facilitate the development and validation of other meth-
odsofanalysisofmagnoorineinotherbiologicalmatrixes
[].

 Evidence-Based Complementary and Alternative Medicine
5.2. Pharmacokinetics of Taspine. Lu et al. () prepared
taspine solid lipid nanoparticles (-SLN) and taspine solid
lipid nanoparticles with galactoside (-GSLN) separately
using the lm evaporation extrusion method. e pharma-
cokinetics and liver target eciency aer IV administrations
of -SLN and -GSLNtoICRmicewerenallycompared
[]. e pharmacokinetics and tissue distribution aer
intravenous administrations of taspine solution and taspine
liposometoICRmicewerecompared.Incorporationinto
liposomes prolonged taspine retention within the systemic
circulation and increased its distribution to the spleen and
liverbutreduceditsdistributiontotheheartandbrain[].
5.3. Pharmacokinetics of Caulophine. Pharmacokinetic stud-
ies have shown that caulophine ()iseasilyabsorbedaer
oral administration, but it is eliminated from the body slowly.
In fact, . h aer treating rats treated with caulophine, the
highest concentration of caulophine was found in the liver.
erefore, hepatic metabolism is probably the main route for
the in vivo processing of caulophine []. Two metabolites
including glucuronide conjugate and N-oxide of caulophine
werefoundinraturineandfecesbyHPLC-MS.Moreover,the
same caulophine glucuronide conjugate was observed in rat
liver microsomes system. However, caulophine glucuronide
conjugatewasnotobservedindoglivermicrosomes[].
6. Cell Membrane Chromatography for
Activity Screening
Cell membrane chromatography(CMC) is a novel bioanity
chromatographic technique. e CMC combined with high
performance liquid chromatography (HPLC) or HPLC/MS
will be of great utility in drug discovery using natural medici-
nal herbs as a source of novel compounds. In reported studies,
the model of CMC in which cell membrane is enriched with
certain receptors is used, as the stationary phase was applied
to screen the target components from medicinal herbs [–
] and to investigate the interactions between drug and
receptor [,]. is system has been successfully applied
to the screening and identication of active components from
C.robustum.
A combined A/CMC-HPLC method was developed
and was successfully applied to recognize, separate, and
identify target components “taspine” and “caulophine” from
C.robustum []. A combined A/CMC with online
HPLC/MS was also established for identifying active com-
ponents from C.robustum acting on human epidermal
growth factor receptor (EGFR) []. Retention fractions on
A/CMC model were captured onto an enrichment column
and the components were directly analyzed by combining a
-portcolumnswitcherwithanLC/MSsystemforsepara-
tion and preliminary identication. Using sorafenib tosylate
as a positive control, taspine () and caulophine ()were
identied as the active molecules which could act on the
EGFR. Other research results showed that taspine ()wasthe
active molecule acting on the tumor vasodilatation [], and
magnoorine () and caulophine ()weretheactive
molecules acting on the human 𝛼1A-adrenoceptor (𝛼1AAR)
[].
is system has been also successfully applied to inves-
tigate the interactions between active compounds from C.
robustum and receptor. A new high-expression vascular
endothelial growth factor receptor- (VEGFR-) CMC
method combined with mathematical treatments was pro-
posed for evaluating taspine-receptor interactions []. A
competitive binding study was performed and the results
indicate that there are multiple types of binding sites on
VEGFR- for taspine (). Following this, Du and coworkers
developed another new high-expression EGFR CMC method
to recognize the ligands acting on EGFR specically and
investigate the anity of getinib/a novel taspine derivative
HMQ to EGFR []. It has been proven that the CMC
method combined zonal elution provides a powerful tech-
nique for the characterization of HMQ binding to the
EGFR.
7. Conclusions and Future Prospects
e present review discusses the chemistry and pharma-
cological aspects of the genus Caulophyllum and especially
provides a detailed analysis of the literature published since
the year of . e state of the science on Caulophyllum
chemistry and pharmacological activity leaves considerable
opportunity for future discoveries.
Two new classes of alkaloids, piperidine-acetophenone
conjugates (–) and uorenone (–)alkaloids,have
been reported from genus Caulophyllum,suggestingthat
piperidine-acetophenone conjugates and uorenone type
alkaloidsareanothertwomajorkindsofmetabolitesthat
existed in this genus Caulophyllum. In addition to common
aglycones (oleanolic acid, hederagenin, echinocystic acid,
and caulophyllogenin), eight other kinds of aglycones have
been found from Caulophyllum species. Diverse aglycones,
monosaccharide residues, and linked modes of sugars are
possible to form diverse structures of triterpene saponins
from genus Caulophyllum.Manynewcompoundshavebeen
identied in recent years, and we are convinced that more
trace constituents with novel structures will be discovered
with the development of new technology for isolation and
identication.
Currently, although many puried compounds have been
tested for activity which are ,,,,,,,,,–,
, and ,only(taspine) is performed in-depth study on its
anti-tumor and anti-angiogenic mechanisms and could serve
as a lead compound in anticancer agent development. Mean-
while, a class of biphenyl derivatives of taspine was designed
and synthesized for screening potential novel anticancer
agents. Besides ,caulophine() was identied as another
active molecule which could act on the EGFR and 𝛼1AAR
by combining 𝛼1AAR/CMC and A/CMC with online
HPLC/MS.  also merits further research to see its action
of mechanisms. On the other hand, a number of compounds
with novel structure skeleton, such as –,–,,,,
,and have previously been isolated, but no further tests
have been performed. It is possible that these compounds are

Evidence-Based Complementary and Alternative Medicine 
usually overlooked due to their low abundance in Caulophyl-
lum. Pharmacokinetic study is also limited for compounds
isolated, mainly involving three active alkaloids ,, and .
So it is very urgent to develop pharmacokinetic study in vivo
for other bioactive compounds in genus Caulophyllum.
Pharmacological studies carried out on crude extracts
and pure metabolites provided pragmatic documents for its
traditional uses and have revealed that this genus is a valuable
source for medicinally important molecules. Many important
biological activities of this genus have been demonstrated
such as anti-inammatory and analgesic eects, antioxidant
eects, antiacetylcholinesterase activity, and antitumor et al.
ough many promising results were conrmed by animal
models, it should be further investigated by clinical trials.
Regarding the constituents contributed to medicinal values,
thendingsindicatedthatalkaloidsandtriterpenesaponins
were regarded as the major constituents in this genus, while
polysaccharides that occurred in the genus are worthy of
further researching their chemical and pharmacological
activities []. However, most of the plant extracts used in the
abovebioassaywerenotwellcharacterized,andthisdefectled
to the diculty to reproduce the reported results. To add the
availability of primary experimental data, suitable analytical
and standardization protocols of plant materials should be
developed, since these are the ground work for convincing
and reproducible pharmacological studies.
e toxicity of Caulophyllum species is not negligible,
mainly involving the teratogenic eects and inducing heart
failureandshockbyingestingbluecohosh.Fromtheviewof
current research results, alkaloid fractions may be responsible
for major toxicity. However, exact individuals are required for
further research by chemical and pharmacological experi-
ments. e future work should be focused on the relationship
between clinical eects and side-eects of Caulophyllum
extracts to screen a safe and eective dosage. Moreover, a
strict quality control procedure should be adopted to guaran-
tee its quality. On the other hand, the alkaloids and triterpene
saponins are two major kinds of constituents in blue cohosh,
which are easily divided by chromatography methods [].
e individual pharmacological tests for fractions of alkaloids
and triterpene saponins should be considered according to
thetraditionalandmodernusesofCaulophyllum plants.
Whether alkaloids and triterpene fractions can be used
separately in the future according to each medical function,
it may be a good choice for Caulophyllum plants for reducing
thedruginteractionandenhancingtheireciency.
Abbreviations
AChE: Antiacetylcholinesterase
AG: Aglycones
AR: Adrenoceptor
Ara: 𝛼-L-Arabinopyranose
Bfgf: Basic broblast growth factor
CMC: Cell membrane chromatography
COX: Cyclooxygenase
CYP: Cytochrome P
DPPH: ,-Diphenyl--picrylhydrazyl
EGFR: Epidermal growth factor receptor
ER: Estrogen receptor
GC/MS: Gas chromatography/mass spectrum
Glc: 𝛽-D-Glucopyranose
HFLPME: Hollow-ber liquid-phase microextraction
HPLC: High performance liquid chromatography
HPLC/MS: High performance liquid
chromatography/mass spectrum
HUVECs: Human umbilical vein endothelial cells
IC50: Half-inhibition concentration
IL: Interleukin
iNOS: Inducible nitric oxide synthase
LPS: Lipopolysaccharide
MIC: Minimal inhibitory concentration
NF-𝜅B: Nuclear factor kappa B
NO: Nitric oxide
NO2: Nitrogen dioxide
PR: Progesterone receptor
REC: Rat embryo culture
Rha: 𝛼-L-Rhamnopyranose
TNF-𝛼: Tumor necrosis factor-𝛼
TV: Tumor vasodilatation
VEGF: Vascular endothelial growth factor
VEGFR-: Vascular endothelial growth factor
receptor-.
Conflict of Interests
e authors declare that they have no conict of interests
regarding the publication of this paper.
Authors’ Contribution
Yong-Gang Xia and Guo-Yu Li equally contributed to this
work.
Acknowledgments
e authors’ work was nancially supported by academic
visitor plans for China Scholarship Council (), the
State Key Creative New Drug Project of th Five-year Plan
of China (ZX), the National Natural Science
Foundation of China (), and New Century Excellent
Talents in Heilongjiang Provincial University.
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