A new aromatic glycoside and its anti-proliferative activities from the leaves of Bergenia purpurascens

Article abstract of DOI:10.1080/14786419.2017.1338278

Chemical investigation of the ethanolic extracts of the dried leaves of Bergenia purpurascens led to the isolation and identification of a new aromatic glycoside, 1-O-β-D-glucopyranosyl-2-methoxy-3-hydroxyl-phenylethene (1), along with other 19 known compounds (2–20). The structure of compound 1 was determined by a detailed analysis using various analytical techniques, including 1D and 2D NMR. In vitro anti-proliferative activities of compound 1 on five human cancer cell lines were evaluated. The results showed that compound 1 possessed the most potent effects with the IC₅₀ values of 14.36?±?1.04?μM against T24 cells. The further bioactivity analysis showed that compound 1 induced apoptosis of T24 cells, and altered anti- and pro-apoptotic proteins, leading to mitochondrial dysfunction and activation of caspase-3 for causing cell apoptosis. The present investigation illustrated compound 1 might be used as a potential antitumour chemotherapy candidate.

Full text of DOI:10.1080/14786419.2017.1338278

Natural Product Research
Formerly Natural Product Letters
ISSN: 1478-6419 (Print) 1478-6427 (Online) Journal homepage: http://www.tandfonline.com/loi/gnpl20
A new aromatic glycoside and its anti-
proliferative activities from the leaves of Bergenia
purpurascens
Shan-Shan Zhang, Zhi-Xin Liao, Ri-Zhen Huang, Cheng-Cheng Gong, Lan-Ju Ji
& Hong-Fa Sun
To cite this article: Shan-Shan Zhang, Zhi-Xin Liao, Ri-Zhen Huang, Cheng-Cheng
Gong, Lan-Ju Ji & Hong-Fa Sun (2017): A new aromatic glycoside and its anti-proliferative
activities from the leaves of Bergenia purpurascens, Natural Product Research, DOI:
10.1080/14786419.2017.1338278
To link to this article: http://dx.doi.org/10.1080/14786419.2017.1338278
View supplementary material
Published online: 12 Jun 2017.
Submit your article to this journal
View related articles
View Crossmark data
Full Terms & Conditions of access and use can be found at
http://www.tandfonline.com/action/journalInformation?journalCode=gnpl20
Download by: [The UC San Diego Library]
Date: 12 June 2017, At: 18:56

Natural Product research, 2017
https://doi.org/10.1080/14786419.2017.1338278
A new aromatic glycoside and its anti-proliferative activities
from the leaves of Bergenia purpurascens
Shan-Shan Zhanga, Zhi-Xin Liaoa, Ri-Zhen Huanga, Cheng-Cheng Gonga, Lan-Ju Jib
and Hong-Fa Sunb
aschool of chemistry and chemical engineering, southeast university, Nanjing, Pr china; bKey laboratory of
tibetan Medicine research, Northwest Institute of Plateau Biology, chinese academy of sciences, Xining, Pr
china
ABSTRACT
ARTICLE HISTORY
received 19 November 2016
accepted 12 May 2017
Chemical investigation of the ethanolic extracts of the dried leaves of
Bergenia purpurascens led to the isolation and identification of a new
aromatic glycoside, 1-O-β-D-glucopyranosyl-2-methoxy-3-hydroxyl-
phenylethene (1), along with other 19 known compounds (2–20).
The structure of compound 1 was determined by a detailed analysis
using various analytical techniques, including 1D and 2D NMR. In vitro
anti-proliferative activities of compound 1 on five human cancer cell
lines were evaluated. The results showed that compound 1 possessed
the most potent effects with the IC50 values of 14.36 1.04 μM against
T24 cells. The further bioactivity analysis showed that compound 1
induced apoptosis of T24 cells, and altered anti- and pro-apoptotic
proteins, leading to mitochondrial dysfunction and activation of
caspase-3 for causing cell apoptosis. The present investigation
illustrated compound 1 might be used as a potential antitumour
chemotherapy candidate.
KEYWORDS
Bergenia purpurascens;
aromatic glycoside; cancer
cell lines; anti-proliferative
activities; docking model
CONTACT Zhi-Xin liao
zxliao@seu.edu.cn
supplemental data for this article can be accessed at https://doi.org/10.1080/14786419.2017.1338278.
© 2017 Informa uK limited, trading as taylor & Francis Group

2
S.-S. ZHANG ET AL.
1. Introduction
The genus Bergenia (Saxifragaceae) includes about 10 species mainly distributed in Central
and East Asia, are evergreen perennial herbs. The rhizomes have been used in herbal formu-
lations to dissolve kidney and bladder stones, and to treat leucorrhoea, pulmonary afflictions
for centuries (Asolkar et al. 1992). Alcohol extracts of the genus Bergenia have been shown
to have analgesic, anti-inflammatory, antibacterial and anti-tussive (Sinha et al. 2001), diuretic
properties (Gehlot et al. 1976), and antilithiatic (Seth et al. 1974; Garimella et al. 2001), anti-
viral (Rajbhandari et al. 2001, 2003). There are many reports of ethnobotanical use of different
Bergenia species for treatment of urinary calculi and other urinary diseases; for treatment of
heart disease, stomach disorders, and anti-diabetic properties (Srivastava et al. 1986).
Furthermore, the genus Bergenia have been discovered to possess antimicrobial, antioxidant
and urease inhibitory activities in recent studies (Agnihotri et al. 2015; Sadat et al. 2015).
Bergenin which was isolated from the genus Bergenia has now been successfully
developed as a listing of drug (Compound Bergenin Tablets) for treating chronic bron-
chitis in China (Zhang 2008). Previous chemical compositions study of the genus Bergenia
mainly focused on the extraction technology, content determination and preparation
of bergenin.
Bergenia purpurascens, one of the main resource of bergenin, grows in moist stone crevices
of alpine distributes inTibet, has long been used as aTibetan medicine for tonic, haemostatic
and anti-tussive actions (Lv & Wang 2003). Systematic studies on the chemical compositions
of this genus were rarely found in literatures, especially for its leaves, so this research aims
to investigate the constitutions of the leaves of B. purpurascens comprehensively; to find
more biological active components for further medicinal application. As a result, a new
compound was obtained from its ethanolic extract which was rarely found in nature, and
was tested for its antitumour and anti-proliferative activities, as well as docking model.
2. Results and discussion
Compound 1 was obtained as white powder, which was analysed for the molecular formula
20
C15H20O8 by HR-ESI-MS (m/z for [M-H]: 327.0736 Calcd 327.1035);ꢀD :+37.88 (c = 0.033, MeOH);
IR (KBr) vmax cm−1: 3227, 1650, 1600, 1509, 1439, 894, 823, 776, 752; UV λmax (CH3OH) (log ε)
230 (1.78), 283 (2.21), 361 (0.23). The 1H NMR spectrum of 1 showed terminal alkenyl protons
at δH 6.97 (1H, dd, J = 6.8, 12.8 Hz, Ha-8), and 6.84(1H, dd, J = 6.8, 8.92 Hz, Hb-8), one alkenyl
proton at δH 6.64 (1H, dd, J = 8.92, 12.8 Hz, H-7), corresponding to two olefinic carbons at
C-8 (δc 118.0) and C-7 (δc 114.8) respectively, with the help of HSQC spectrum. The position
of the double bond group was assigned at C-5 (δc 151.9) due to the proton signal of H-7 (δH
6.64) associated with C-4 (δc 115.9), C-6 (δc 118.1), and the proton signals of Ha-8, Hb-8 (δH
6.97 and 6.84) relating to C-5 (δc 151.9) from the HMBC. According to δH 3.69 (3H, s) and δc
55.8, deduced that there was a methoxy group, and it correlated with C-2 (δc 152.6) from
HMBC spectrum. In the low field of 1H NMR, one proton at δH 9.0 (1H, s), it was the character
of hydroxyl group, and the hydroxyl proton signals was associated with C-2 (δc 152.6) and
C-4 (δc 115.9) from the HMBC analysis, so the hydroxyl group was assigned at C-3 (δc 154.7).
Key HMBC correlations were shown in Figure 1.
Compound 1 was hydrolysed with dilute hydrochloric acid, paperTLC showed the Rf value
of sugar moiety was equal to the glucose standard sample. The terminal proton signal of
glucose H-1′ δH 4.63 (1H, d, J = 7.6 Hz) could judge as β-configuration and it was connected

NATURAL PRODUCT RESEARCH
3
to C-1 (δc 150.8) from the HMBC. In theTOCSY spectrum, from the glucose terminal hydrogen
(δH 4.63), this sugar spin-coupled system was got, which could be attributed other hydrogen
protons, 4.63-3.12-3.15-3.23-3.25-3.45 (glucose). Hence, the structure of new compound 1
was confirmed as 1-O-β-D-glucopyranosyl-2-methoxy-3-hydroxyl-phenylethene.
The known compounds (2–20) were ursolic acid (2) (Ibrahim et al. 2014), 3β,5α-
dihydroxy-15-cinnamoyloxy-14-oxolathyra-6Z,12E-diene (3) (Wang et al. 2008), oleanolic
acid (4) (Guo et al. 2008), 2α-hydroxyursolic acid (5) (Seo et al. 1975), hydroquinone (6)
(Song et al. 2007), gallic acid (7) (Deng & Qin 2008), Ferulic Acid (8) (Choi et al. 2005),
11-O-galloylbergenin (9) (Zhang et al. 2007), 11-O-(4′-hydroxybenzoyl) bergenin (10)
(Kashima & Miyazawa 2012), ardimerin (11) (Ryu et al. 2002), taraxerol (12) (Feng et al. 2008),
β-sitosterol (13) (Shen et al. 2012), betulinic acid (14) (Zhang, Li et al. 2014), ocimol (15)
(Siddiqui et al. 2007), kaempferol (16) (Kuroyanagi et al. 1978), bergenin (17) (Chen & Yang
2008), daucosterol (18) (Kim et al. 1996), β-sitosterol palmitate (19) (Wei et al. 2013), arbutin
(20) (Sun et al. 2006). All the above compounds were identified by the spectral data (UV, IR,
ESI-MS, 1H and 13C NMR) with those reported in the literature.
2.1. Cytotoxic effects of compounds
Seven compounds against the five cell lines (MGC-803, T24, HepG2, NCI-H460 and HL-7702)
were evaluated with MTT assay (Table 1). Compared with other compounds, 1 showed good
Figure 1. Key hMBc correlations of the new compound 1.
Table 1. cell growth inhibition activity of compounds.
Compound
IC50 (μM)
NCI-H460
MGC-803
HepG2
T24
HL-7702
1
18.10 1.15
27.95 1.42
18.81 1.04
35.16 0.78
>100
83.56 0.98
26.95 1.12
5.06 1.21
20.93 1.07
42.78 1.44
26.42 0.87
>100
>100
>100
60.15 1.28
5.03 0.85
17.69 1.17
53.89 1.36
25.68 1.04
>100
>100
>100
36.47 0.93
38.55 1.16
14.36 1.04
48.15 1.09
46.54 1.23
31.29 1.35
>100
56.72 0.84
26.95 1.35
4.37 0.57
>100
>100
>100
>100
>100
>100
>100
3
9
10
11
15
17
hcPta
69.47 1.84
aused as positive control.

4
S.-S. ZHANG ET AL.
inhibitory activity, especially for cancer cell lines NCI-H460 and T24, reduced NCI-H460 cells
percentages significantly, with IC50 values of 17.69 μM. Furthermore, compound 1 showed
the strongest inhibitory effect on the growth of T24 cells (IC50 = 14.36 μM). However, com-
pound 1 barely inhibited the growth of HL-7702, which is a normal cell line, at concentrations
up to 100 μM.
2.2. Morphological characterisation of cell apoptosis of T24 cells by Hoechst
33258
In order to further validate the cell apoptosis upon treatment of compound 1, T24 cells were
stained with Hoechst 33258 staining after the treatment for 24 h at different concentrations
(0, 15, 30 μM). Our experimental observation showed that, in the control group, most of the
cells exhibited the weak blue fluorescence of normal cells (Figure S1). After the treatment
of compound 1, some cells emitted brilliant blue fluorescence, and nuclei of more T24 cells
appeared hyper condensed (brightly stained). Remarkably, the numbers of apoptotic nuclei
containing condensed chromatin increased significantly after the T24 cells were treated with
compound 1 for 24 h, indicating that apoptosis of the T24 cells was induced by compound
1 in a concentration-dependent manner.
2.3. Effects of compound 1 on the induction of apoptosis
In order to confirm whether compound 1 induced reduction in cell viability due to the
induction of apoptosis, the apoptotic rates of T24 cells treated with compound 1 at various
concentrations of 0, 15, 30 μM (Figure S2). As shown in Figure S2, few (5.46%) apoptotic cells
were present in the control panel, in contrast, the population rose to 17.99% at the concen-
tration of 15 μM after treatment with compound 1 for 24 h. Further increase to 49.8%
occurred after treatment with 1 at the concentration of 30 μM. These results clearly confirmed
that compound 1 could cause a notable increase of cellular apoptosis in dose-dependent
manner from 0 to 30 μM.
2.4. Compound 1 induced expression of pro- and anti-apoptotic proteins
The Bcl-2 family members are important regulators of the mitochondrial apoptotic pathway.
Two most important members of Bcl-2 family, the anti-apoptotic protein Bcl-2 and the
pro-apoptotic protein Bax, are key regulators of this progress (Zhang, Nie et al. 2014). The
effects of compound 1 on the constitutive levels of Bax, Bcl-2 and caspase-3 in T24 cells are
given in Figure S3, compared with the control group, 1 induced a significant increase of Bax
level and an inhibition on the expression of Bcl-2, in a dose-dependent manner.
2.5. Docking model
As biological activity studies suggested that compound 1 could dramatically inhibit the
expression of Bcl-2 (Figure S3), compound 1 was docked into the available crystal structure
of Bcl-2 (PDB: 1bxl) in order to rationalise the observation. The binding modes of compound
1 in the binding site of Bcl-2 are depicted in Figure 2. The Surflex docking scores is 9.00 for
1, where higher scores indicate greater binding affinity. Figure 2 shows that the interacting

NATURAL PRODUCT RESEARCH
5
Figure 2. Molecular modelling of compound 1 in complex with Bcl-2.
mode of compound 1 in the binding site is surrounded by Arg6, Glu7, Val10, Ser14, Trp24,
Asp29, Arg34 and Thr35. In particular, compound 1 forms some hydrogen bonds with the
polar amino acids Glu7, Asp29, Arg34 and Thr35, respectively, suggesting a probable strong
electrostatic interaction with the protein. Molecular docking analysis showed that compound
1 has inhibition effect on expression of Bcl-2 protein may be related with the binding to
Bcl-2.
3. Experimental
3.1. General methods
1H and 13C NMR spectra were measured on a Bruker Avance DRX-400 spectrometer. IR spectra
were recorded on a NICOLET IR200 FT-IR spectrophotometer. HR-ESI-MS was carried out on
an AgilentTechnologies 6224TOF LC-MS apparatus. TLC was conducted on silica gel HSGF254
plates (10–40 μm; Yan tai Jiang you Chemical, Inc.). Column chromatography (CC) was per-
formed with silica gel (200–300 mesh, Qingdao Marine Chemical, Inc.), MCI Gel (CHP 20P,
75–150um, Mitsubishi Kasei Corporation).
3.2. Plant material
The leaves of B. purpurascens were collected in September 2015 from Jiuzhi County, Guoluo
City, Qinghai Province, China, and the plant was identified by Prof. Zhu-Nan Gong of College
of Life Science, Nanjing Normal University, Nanjing, China. A voucher specimen (No. 15-09-
01-01) was deposited at the Natural Products Chemistry Laboratory of Southeast University,
Nanjing, China.
3.3. Extraction and isolation
The leaves of B. purpurascens (5.0 kg) were extracted with 95% ethanol (30.0 L × 5) at
room temperature. After removal of the solvent, the extractum (327.0 g) was suspended
in water (3000 mL), partitioned sequentially with petroleum ether (PE) (2000 mL × 5
times), EtOAc (2000 mL × 5 times) to yield crude PE (100.3 g), EtOAc (183.3 g) extracts
respectively. The crude PE extract was decolorised on MCI GEL, then a sample (19.0 g)

6
S.-S. ZHANG ET AL.
was subjected to silica gel CC using a gradient system with increasing polarity of PE/
EtOAc (from 50:1 to 1:6, v/v) to yield 12 (18 mg), 13 (347 mg), 14 (165 mg), 15 (12 mg),
16 (23 mg), 17 (1003 mg), 18 (368 mg), 19 (16 mg), 20 (27 mg). The EtOAc extract decol-
orized on MCI GEL (90.0 g), then used the same method to afford 1 (610 mg), 2 (338 mg),
3 (10 mg), 4 (18 mg), 5 (23 mg), 6 (189 mg), 7 (207 mg), 8(20 mg), 9 (903 mg), 10 (486 mg),
11 (11 mg).
3.4. Docking model
All the docking studies were carried out using Sybyl-X 2.0 on a windows workstation. The
crystal structure of the tubulin in complex with colchicine was retrieved from the RCSB
Protein Data Bank (PDB: 1bxl.pdb) (Enyedy et al. 2001). The 3D structures of these selected
compounds were first built using Sybyl-X 2.0 sketch followed by energy minimisation using
the MMFF94 force field and Gasteiger-Marsili charges. We employed Powell’s method for
optimising the geometry with a distance dependent dielectric constant and a termination
energy gradient of 0.005 kcal/mol. All the selected compounds were automatically docked
into the colchicine binding pocket of Bcl-2 by an empirical scoring function and a patented
search engine in the Surflex docking program. The polar hydrogen atoms were added and
also the automated docking manner was applied in the present work. Other parameters
were established by default to estimate the binding affinity characterised by the Surflex-
Dock scores in the software. A higher score represents stronger binding affinity. The optimal
binding pose of the docked compounds was selected based on the Surflex scores and visual
inspection of the docked complexes.
3.5. 1-O-β-D-glucopyranosyl-2-methoxy-3-hydroxyl-phenylethene
20
White powder; m.p.145–147°C; ꢀD :+37.88°(c = 0.033, MeOH); IR (KBr) vmax cm−1: 3227, 1650,
1600, 1509, 1439, 894, 823, 776, 752; UV λmax (CH3OH) (log ε) 230 (1.78), 283 (2.21), 361 (0.23);
1
HR-ESI-MS: m/z 327.0736 [M-H]− (C15H20O8 Calcd 327.1035). H NMR (400 MHz, DMSO) δH
(ppm, J Hz): 9.0 (1H, s, OH-3), 6.97 (1H, dd, J = 6.8, 12.8 Hz, Ha-8), 6.84 (1H, dd, J = 6.8, 8.92 Hz,
Hb-8), 6.66 (1H, d, J = 2.0 Hz, H-4), 6.86 (1H, d, J = 3.2 Hz, H-6), 6.64 (1H, dd, J = 8.92, 12.8 Hz,
H-7), 3.69 (3H, s, –OCH3), 4.63 (1H, d, J = 7.6 Hz, H-1′), 4.55 (1H, t, J = 5.6 Hz, OH-6′), 4.99 (1H,
d, J = 5.2 Hz, OH-4′), 5.05 (1H, d, J = 4.4 Hz, OH-3′), 5.25 (1H, d, J = 4.8 Hz, OH-2′), 3.45 (1H, m,
H-6′), 3.25 (1H, m, H-3′), 3.23 (1H, m, H-5′), 3.15 (1H, m, H-2′), 3.12 (1H, m, H-4′); 13C NMR
(100 MHz, DMSO) δc (ppm): 55.8 (–OCH3), 61.2 (C-6′), 70.2 (C-4′), 73.7 (C-2′), 77.1 (C-5′), 77.4
(C-3′), 102.2 (C-1′), 114.8 (C-7), 115.9 (C-4), 118.0 (C-8), 118.1 (C-6), 150.8 (C-1), 151.9 (C-5),
152.6 (C-2), 154.7(C-3).
4. Conclusions
A new aromatic glycoside (1), and 19 known compounds (2–20), were isolated from the
leaves of B. purpurascens. The structure of the new compound was identified on the basis of
spectroscopic analysis. The novel compound demonstrated potent cytotoxic activities
against the T24 cancer cell lines. The further bioactivity analysis showed that compound 1
induced apoptosis of T24 cell lines.

NATURAL PRODUCT RESEARCH
7
Supplementary material
The biological experiments and original spectra of NMR, HR-ESI-MS data of the new compound (1) are
available as Supporting Information.
Acknowledgment
We are grateful to Prof. Zhu-Nan Gong of College of Life Science, Nanjing Normal University, Nanjing,
China, for the identification of the plant material.
Disclosure statement
No potential conflict of interest was reported by the authors.
Funding
The project was funded by State Key Laboratory for Chemistry and Molecular Engineering of Medicinal
Resources (Guangxi Normal University) [grant number CMEMR 2016-B06].
References
Agnihotri V, Sati P, Jantwal A, Pandey A. 2015. Antimicrobial and antioxidant phytochemicals in leaf
extracts of Bergenia ligulata: a Himalayan herb of medicinal value. Nat Pro Res. 29:1074–1077.
Asolkar LV, Kakkar KK, Chakre OJ. 1992. Glossary of Indian medicinal plants with active principles. New
Delhi: PID, CSIR; p. 122.
Chen P, Yang JS. 2008. Chemical constituents from the Caesalipinia millettii. China J Chin Mater Med.
33:1852–1855.
Choi BR, Hong SS, Han XH, Hwang JS, Lee MH, Hur JD, Hwang BY, Ro JS. 2005. Antioxidant constituents
from Portulaca oleracea. Nat Prod Sci. 11:229–232.
Deng AJ, Qin HL. 2008. Studies on chemical constituents of fruits of Bridelia tomentosa. China J Chin
Mater Med. 33:158–160.
Enyedy IJ, Ling Y., Nacro K, Tomita Y, Wu X, Cao, Y, Guo R, Li B, Zhu X, Huang Y, Long YQ. 2001. Discovery
of small-molecule inhibitors of Bcl-2 through structure-based computer screening. J Med Chem.
44:4313–4324.
Feng B, Gong XJ, Shi LY, Jian G, Pei YH, Wang YQ. 2008. Studis on phenolic compounds from Stellera
chamaejasme. China J Chin Mater Med. 33:405–408.
Garimella TS, Jolly CI, Narayanan S. 2001. In vitro studies on antilithiatic activity of seeds of Dolichos
biflorus Linn and rhizomes of Bergenia ligulata Wall. Phytother Res. 15:351–355.
Gehlot NK, Sharma VN, Vyas DS. 1976. Some pharmacological studies on ethanolic extract of roots of
Bergenia ligulata. Indian J Pharmacol. 8:92–94.
Guo N, Chen XQ, Zhao QS. 2008. A new polyisoprenylated benzoylphloroglucinol derivative from
Hypericum henryi subsp. uraloides (Guttiferae). Acta Bot Yunnan. 30: 515–518.
Ibrahim M, Ambreen S, et al. 2014. Phytochemical investigation on Eucalyptus globulus Labill. Asian J
Chem. 26:1011–1014.
KashimaY, Miyazawa M. 2012. Synthesis and biological evaluation of bergenin analogues as mushroom
tyrosinase inhibitors. Arch Pharmacal Res. 35:1533–1541.
Kim DK, Kwak JH, Song KW, Kwon HC, Zee OP, Lee KR. 1996. Phytochemical constituents from A. conitum
pseudolaeve Var. erectum. Saengyak Hakhoechi. 27:75–79.
Kuroyanagi M, Fukuoka M, Yoshihira K, Natori S, Yamasaki K. 1978. Confirmation of the structure of
tiliroside, an acylated kaempferol glycoside, by 13C-Nuclearmagnetic resonance. Chem Pharm Bull.
26:3594–3596.

8
S.-S. ZHANG ET AL.
Lv XM, Wang JX. 2003. Research progress in genus Bergenia plants. Chin Med Mat. 26:58–60.
Rajbhandari M, Wegner U, Jülich M, Schöpke T, Mentel R. 2001. Screening of Nepalese medicinal plants
for antiviral activity. J Ethnopharmaco. 74:251–255.
Rajbhandari M, Wegner U, Schopke T, Lindequist U, Mentel R. 2003. Inhibitory effect of Bergenia ligulata
on influenza virus A. Pharmazie. 58:268–271.
Ryu GS, Lee SY, Kim BS, Ryu SY, Hwang HJ, Choi BW, Lee BH, Jung DS. 2002. Ardimerin, a new dimeric
lactone from the herb of Ardisia japonica. Nat Prod Sci. 8:108–110.
Sadat A, Uddin G, Alam M, Ahmad A, Siddiqui BS. 2015. Structure activity relationship of bergenin,
p-hydroxybenzoyl bergenin, 11-O-galloylbergenin as potent antioxidant and urease inhibitor
isolated from Bergenia ligulata. Nat Pro Res. 29:2291–2294.
Seo S, Tomita Y, Tori K. 1975. Biosynthesis of ursene-type triterpenes from sodium [1,2-13C] acetate in
tissue cultures of Isodon japonicushara and reassignments of 13C NMR signals in urs-12-enes. J Chem
Soc Chem Commun. 1:954–955.
Seth SDS, Prabhakar MC, Bapna BC, Arora RB. 1974. Studies on the antilithiatic property of Berginia
ligualata. Res Indian Med. 9:1–3.
Shen L, Jiang SP, Zhu HJ. 2012. Chemical constituents in herb of Incravillea younghusbandii Sprague.
Nat Prod Res Dev. 24:1210–1213.
Siddiqui BS, Aslam H, Ali ST, Begum S, Khatoon N. 2007. Two new triterpenoids and a steroidal glycoside
from the aerial parts of Ocimum basilicum. Chem Pharm Bull. 55:516–519.
Sinha S, Murugesan T, Maiti K, Gayen JR, et al. 2001. Antibacterial activity of Bergenia ciliata rhizome.
Fitoterapia. 72:550–552.
Song YL, Feng ZB, Cheng YX, Gao JM. 2007. Chemical components of Chaenomeles speciosa (Sweet)
Nakai. Acta Bot Boreal Occident Sin. 27:0831–0833.
Srivastava TN, Rajasekhren S, Badola DP, Shah DC. 1986. An index of the available medicinal plants,
used in Indian system of medicine from Jammu and Kashmir State. Ancient Sci Life. 6:49–63.
Sun XH, Shen GM, Tian X. 2006. Chemical components of Scrofella Chinensis. Acta Bot Boreal Occident
Sin. 26:0412–0415.
Wang H, Zhang XF, Zhou Y, Peng SL, Zhou DW, Ding LS. 2008. Two new diterpenes from Euphorbia
kansuensis. Fitoterapia. 79:262–266.
Wei XX, Li YT, Jin YQ. 2013. Studies on chemical constituents from Husks of Tatary Buckwheat. Nat Prod
Res Dev. 25:44–46.
Zhang N, Li N, SunYN, Li JL, Xing SS, Tuo ZD, Cui L. 2014. Diacylglycerol compounds from Barks of Betula
platyphyllawith inhibitory activity against acyltransferase. Chin Herb Med. 6:164–167.
Zhang SS, Nie SP, Huang DF, Feng YL, Xie MY. 2014. A novel polysaccharide from Ganoderma atrum
exerts antitumor activity by activating mitochondria-mediated apoptotic pathway and boosting
the immune system. J Agric Food Chem. 62:1581–1589.
ZhangYH. 2008. Determination of bergeninum and chlorphenamine maleate in compound bergeninum
tablets by HPLC. China Pharmaceuticals. 17:25–26.
Zhang W, Xu KP, Zeng GY, Tan JB, Li FS, Zhou YJ, Tan GS. 2007. Studies on chemical constituents of
Ardisiabicolor Walker. Chin Pharm J. 42:656–658.