July 2014
Regular Article
Chem. Pharm. Bull. 62(7) 695–699 (2014)
695
Hepatoprotective Triterpenoid Saponins from Callicarpa nudiflora
,a
,c
Bo Huang,a,b,# Hui-Zheng Fu,a,# Wei-Kang Chen,a Yue-Hua Luo,* and Shuang-Cheng Ma*
a Jiangxi Provincial Institute for Drug and Food Control, Jiangxi Provincial Engineering Research Center for
b
Drug and Medical Device Quality; Nanchang 330029, China: Pharmaceutical Department of Nanchang University;
c
Nanchang 330006, China: and National Institutes for Food and Drug Control; Beijing 100050, China.
Received February 17, 2014; accepted April 1, 2014; advance publication released online May 8, 2014
Four new triterpenoid saponins, 2α,3α,19α,24-tetrahydroxyolean-12-en-28-oic-acid 28-O-β-D-gluco-
pyranosyl ester (1), 2α,3α,19α,23-tetrahydroxyolean-12-en-28-oic-acid 28-O-β-D-xylopyranosyl-(1→2)-β-D-glu-
copyranosyl ester (2), 2α,3α,19α-trihydroxyolean-12-en-28-oic-acid 28-O-β-D-xylopyranosyl-(1→2)-β-D-gluco-
pyranosyl ester (3), 2α,3α,23,29-tetrahydroxyurs-12,19-dien-28-oic-acid 28-O-β-D-glucopyranosyl ester (4),
together with three known compounds (5–7), were isolated from the leaves of Callicarpa nudiflora HOOK.
Their structures were established by means of spectroscopic methods and chemical evidence. Hepatoprotec-
tive activities of the isolated compounds against D-galactosamine-induced toxicity have been tested. Among
them, compounds 1–3 showed pronounced hepatoprotective activities against D-galactosamine-induced toxic-
ity in WB-F344 rat hepatic epithelial stem-like cells.
Key words Callicarpa nudiflora; triterpenoid saponin; hepatoprotective activity
Callicarpa nudiflora HOOK is distributed widely in Guang- (Table 1). The 13C-NMR spectrum of 1 displayed 36 carbon
dong, Guangxi, and Hainan Provinces in China. The plant is signals, of which 6 were attributed to the sugar moiety and the
used as traditional Chinese herbal medicines for the treatment remaining 30 to the aglycone. Signals at δC 124.3, 144.8, and
of inflammation and bleeding.1) Previous investigation on C. 177.7 were assigned to a pair of typical olefinic carbons and
nudiflora has led to the isolation of iridoids, flavonoids, trit- a carboxy carbonyl carbon, respectively. These spectroscopic
erpenoides, and phenylpropanoid glycosides.2–5) Some of them data indicated that 1 is an oleanene-type triterpene8) with four
have been shown to exhibit anti-inflammatory, antibacterial, hydroxyl groups, a trisubstituted double bond. The two oxy-
cytotoxic and hemostatic activies.6) Our previous phytochemi- methylene proton signals at δH 3.87 and 4.15, which correlated
cal study of C. nudiflora yielded two triterpenoid glycosides, with the carbon resonance at δC 65.7 in the HSQC spectrum,
an acylated flavone glycoside, and three furofuran lignans.6,7) showed heteronuclear multiple bond connectivity (HMBC)
Further investigation on the 80% EtOH extract of this plant correlations with C-3, C-4, and C-24, justifying its assignment
resulted in the isolation of four new compounds, 2α,3α,19α, to C-23. The assignment of hydroxyl groups at C-2 and C-3
24-tetrahydroxyolean-12-en-28-oic-acid 28-O-β-D-glucopyra- was confirmed from the HMBC correlations from H-2 to C-4
nosyl ester (1), 2α,3α,19α,23-tetrahydroxyolean-12-en-28-oic- and C-10 and from H-1, H-2, H-23, and H-24 to C-3. Com-
acid 28-O-β-D-xylopyranosyl-(1→2)-β-D-glucopyranosyl ester parison of the NMR spectroscopic data of 1 with sericoside9)
(2), 2α,3α,19α-trihydroxyolean-12-en-28-oic-acid 28-O-β-D- demonstrated that two compounds were almost identical
xylopyranosyl-(1→2)-β-D-glucopyranosyl ester (3), 2α,3α,23, except for the A ring. The splitting pattern of H-3 (1H, d,
29-tetrahydroxyurs-12,19-dien-28-oic-acid 28-O-β-D-glucopy- J=1.8Hz) suggested that 1 was the C-3 epimer of sericoside.
ranosyl ester (4), along with three known compounds, luteolin Thus, the C-3 hydroxy group of 1 was α-orientation. From the
(5), 5,4′-dihydroxyl-3,7,3′-trimethoxyflavone (6), caffeic acid foregoing evidences it was concluded that 1 was a glycoside
(7). Reported herein are the isolation, structure elucidation of tetrahydroxyolean-12-en-28-oic acid. Acid hydrolysis of
and biological activity of these compounds.
1 with 2M HCl afforded D-glucose, which was identified by
GC analysis of its trimethylsilyl L-cysteine derivatives.10) In
the HMBC spectrum of 1, the presence of correlations from
Results and Discussion
Compound 1 was obtained as a white powder. The mo- Glc-H-1 (δH 6.38) to C-28 (δC 177.7) confirmed that the glu-
lecular formula, C36H58O11, was determined by high reso- cose unit is located at C-28 of the aglycone. The α-orientation
lution-electrospray ionization-mass spectra (HR-ESI-MS) of 2-OH, 3-OH, and 19-OH in 1 was deduced by analysis of
(m/z 665.3878 [M−H]−, Calcd for 665.3901). The 1H-NMR the nuclear Overhauser effect spectroscopy (NOESY) spec-
spectrum of 1 in pyridine-d5 showed six methyl singlets (δH trum which showed nuclear Overhauser effect (NOE) correla-
0.99, 1.09, 1.15, 1.19, 1.55, 1.68). Additional proton resonances tions between the following proton pairs: H-2/H-3, H-2/H-24,
observed included those ascribed to an olefinic proton at δH H-2/H-25, and H-19/H-30. Thus, compound 1 was elucidated
5.52 (1H, brs), two oxymethine protons at δH 4.48 (1H, m) as 2α,3α,19α,24-tetrahydroxyolean-12-en-28-oic-acid 28-O-β-
and 4.62 (1H, d, J=1.8Hz), two oxymethylene protons at δH D-glucopyranosyl ester.
3.87 and 4.15 (1H each, d, J=10.8Hz), and an anomeric proton
Compound 2 had the molecular formula C41H66O15, as de-
signal at δH 6.38 (1H d, J=8.4Hz) correlated in the hetero- duced from the HR-ESI-MS (m/z 821.4290 [M+Na]+, Calcd
nuclear single quantum coherence (HSQC) spectrum with for 821.4294). The 1H-NMR spectrum of 2 was similar to
an anomeric carbons at δC 96.4 in the 13C-NMR spectrum those of 1 except for the configuration of C-4 and an ad-
ditional set of β-D-xylose resonances. The proton signal at
δH 0.71 assigned to H-24 of 2 was shifted by −0.97ppm
compared to that of 1, suggesting that the aglycone of 2 was
The authors declare no conflict of interest.
#These authors contributed equally to this work.
© 2014 The Pharmaceutical Society of Japan
*To whom correspondence should be addressed. e-mail: emailluo@sohu.com;