Content uploaded by Gina M Méndez-Callejas
Author content
All content in this area was uploaded by Gina M Méndez-Callejas on Sep 26, 2018
Content may be subject to copyright.
August 30, 2018
Archives • 2018 • vol.2 • 113-122
http://pharmacologyonline.silae.it
ISSN: 1827-8620
ANTIPROLIFERATIVE ACTIVITY OF EXTRACTS OF GNAPHALIUM GRACILE H.B.K. AGAINST
CANCER CELL LINES
Torrenegra-Guerrero, R. D. 1*; Rodriguez-Mayusa, J. 1; Mendez-Callejas, G. M.2; Canter, R. 3;
Whitted, C. 3 and Palau, V. E. 3,4
1Facultad de Ciencias, Universidad de Ciencias Aplicadas y Ambientales Bogotá, Colombia
2Facultad de Ciencias de la Salud, Universidad de Ciencias Aplicadas y Ambientales, Bogotá, Colombia
3Department of Pharmaceutical Sciences, Bill Gatton College of Pharmacy, 4Department of Internal
Medicine, Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614.
*rtorrenegra@udca.edu.co
Abstract
Ethanol and n-hexane extracts obtained from the leaves and inflorescences of Gnaphalium gracile,
were tested at different concentrations to evaluate their antineoplastic activities on pancreatic, colon,
and prostate cancer cell lines by examining mitochondrial function. The polar extracts of both, leaves
and inflorescences which contain gnaphalin, quercetin, and 3-methoxy quercetin, exhibited cytotoxicity
against every cell line tested with EC50 values ranging between 20.23±1.185 µg/mL and 70.71±1.1419
µg/mL.
The most remarkable values were observed in pancreatic cancer Panc 28 and androgen-dependent
prostate LnCaP cells, with EC50 values of 20.23±1.185 and ˂25µg/mL, and androgen-independent
prostate cancer PC-3, colon HCT-116 and pancreatic MIA PaCa cells with values ranging between
28.84±1.1766 and 34.41±1.057 µg/mL. The non-polar extract derived from leaves demonstrated
significant cytotoxicity towards colon cancer HCT-116 cells, with an EC50 of 39.46±1.0617 µg/mL.
However, the non-polar extract from the inflorescences did not have an appreciable effect on cell
proliferation of any of the cell lines tested except for androgen-independent prostate cancer PC-3 cells
with an EC50 of 62.05±1.237 µg/mL. The data obtained support the traditional use of G. gracile and
suggest the polar extracts from aerial parts, as an interesting source for the development of novel
antineoplastic agents.
Keywords: Gnaphalium gracile, anti-neoplastic activity, pancreatic cancer, colon cancer, prostate cancer
PhOL Torrenegra-Guerrero, et al. 114 (pag 113-122)
http://pharmacologyonline.silae.it
ISSN: 1827-8620
Introduction
Gnaphalium gracile H.B.K is a species that belongs
to the family Asteraceae. In equatorial regions it
grows between 2000 and 3200 meters above sea
level and it has broad synonymy [1]. The genus
Gnaphalium has worldwide distribution, with the
greatest diversity of species found in the Andean
regions of South America. In Colombia, these
species grow in association with plants belonging to
the genus Achyrocline with which they share
morphological similarities that may lead to
identification errors. Plants from these genera also
produce similar flavonoids that may confer closely
related medicinal properties to species that in turn,
farmers use interchangeably for the same medicinal
purposes and commonly name vira-vira. Species of
the Gnaphalium genus are advocated in the
treatment of several different diseases, such as, skin
infections, bronchial disorders, inflammation, and
cancer in different organs [2]. Gnaphalium elegans is
popularly used against prostate cancer as well as
Gamochaeta purpureum, a plant also known as vira-
vira. Thus, species of several genera belonging to
the tribe Inuleae, such as Gnaphalium, Achyrocline
and Conyza, are sold in the popular markets of
medicinal plants as vira-vira, to treat the same
illnesses because they have similar medicinal
effects. These properties have been attributed to
flavonoids produced as active metabolites that may
convey these healing properties. A number of
studies have sought to identify the secondary
metabolites produced by different species of
Gnaphalium [3-7]. These reports include the
descriptions of flavonoids as diterpenes [8] with an
unsubstituted ring B, and methoxylation on various
carbons of rings A and C [9-11]. In a recent study, it
was found that the minor variations of ring
substitution of flavonoid isomers lead to
preferential binding to different DNA sequences [11]
and specific cytotoxic actions on cancer cell lines
with varying differentiation status [12]. A study of
the chemistry of the species Gnaphalium gracile
H.B.K. reports the presence of methoxylated
flavonoids, gnaphaline, and others flavonoids with
various hydroxylations in their rings, 3-methoxy
quercetin and quercetin [13]. Because active
metabolites may occur in different parts of the
plant, we hypothesized that extracts obtained with
solvents of different polarity from leaves or
inflorescences may have differential anticancer
properties and effects on cancer cell lines of various
carcinomas. In this study we present the cytotoxic
effects of polar and apolar extracts from leaves and
inflorescences of G. gracile H.B.K. on pancreatic,
colon, and prostate cancer cells with varying
tumorigenic and differentiation status, and
determine the plant sources that may lend support
to its traditional use in the treatment of cancer.
Methods
Procedure to collect and process the plant
material.
Plants of G. gracile H.B.K. were collected during
the months of July through September in the area
surrounding the Tominé reservoir, in the
municipality of Guatavita, in Cundinamarca,
Colombia. The specimens were identified at the
Colombian National Herbarium. Leaves and
inflorescences were separated, dried in the shade,
and soxhlet extracted with AcOEt. The use of the
latter solvent, avoids the extraction of salts and
sugars that may interfere with the extraction of the
fractions of interest. From 230g of inflorescences is
avoided 17g of extract was obtained. Following the
same procedure, but separately, 70g of extract was
obtained from 645g of leaves and stems. 5g of each
extract were fractionated S/L sequentially with
petrol and ethanol, to obtain the apolar fractions (in
petrol) and polar (in EtOH) of leaves and
inflorescences. The fractions were designated as
EFapolar, EFpolar, and EHpolar EHapolar for
inflorescences and leaves respectively, and were
subsequently dried under vacuum. A 10g sample of
leaf extract was subjected to column
chromatography on SiGel, with mobile phase Petrol:
EtOAc 8: 2. Fractions of 50 mL were collected and
isolated and identified as stigmasterol,
dehidroestigmasterol and gnaphaline in fractions 6
and 12 respectively, the compounds were purified by
crystallization in hexane. In the same manner,
gnaphalin was identified after isolation from
inflorescence extracts. It was found in a higher
proportion of about 0.3% of dry material. Column
chromatography with mobile phase RP18 and EtOH :
water 7:3, using the polar extract of leaves or
inflorescences, yielded quercetine, and a mixture of
3- methoxyquercetin and quercetin. The compounds
PhOL Torrenegra-Guerrero, et al. 115 (pag 113-122)
http://pharmacologyonline.silae.it
ISSN: 1827-8620
were identified by comparing their HNMR spectra
with those of pure samples of gnaphaline, 3-
methoxyquercetin and quercetin (figure 1).
Cell Lines and Culture Conditions
Tumor derived cells that originated in multiple
tissue sites, including: colon (Caco-2 and HCT-116),
pancreas (MIA PaCa), and prostate (LNCaP and PC-
3), were obtained from the American Tissue Type
Culture Collection (Manasas, VA), and maintained
according to the supplier instructions. The Panc-28
cell line was a gift from Dr. Paul Chiao, at the
University of Texas M. D. Anderson Cancer Center
[14] and was grown in tissue culture in the same
manner as pancreatic cell line MIA PaCa-2, in
Dulbecco’s modified Eagle’s medium with high
glucose, supplemented with 10% serum and
penicillin/streptomycin (GIBCO, Life Technologies,
Carlsbad, CA). All cells were seeded in 48 well plates
and allowed to reach 75% confluency before
treatment.
Cell Viability Assay
Cells were treated with either vehicle or (EH)
polar, (EH) apolar, (EF) polar, (EF) apolar G. gracile
extracts from leaves and inflorescences
respectively, at concentrations of 25, 50, or 125
µg/mL. The dissolution vehicle was dimethyl
sulfoxide at a maximum final concentration of 0.4%
in the treated well (Sigma- Aldrich, St. Louis, MO).
After 24 hours of incubation, 3-(4, 5-methyl-thiazol-2-
yl)-2, 5-diphenyl-tetrazolium bromide (MTT) (Sigma-
Aldrich) was added at 500ug/mL/well for 3 hours.
Formazan products were solubilized with acidified 2-
propanol (0.1N HCl) and 0.1% NP-40. Assays were
quantified by reading optical density at a
wavelength of 590 nm using a Biotek PowerWave
XS2 plate reader (Winooski, VT). Statistical analyses
where done in an IBM SPSS statistics 20, and the
inhibitory concentration EC50 value was defined as
the extract concentration which caused a 50%
decrease in the cell viability of the control. Data
from assays displaying decrease of cell viability
≥50%, were evaluated by nonlinear regression
analysis (GraphPad Prism, La Jolla, CA), and
represented as the effective concentration required
to decrease 50% of cell viability (EC50).
Results
The polar extract derived from the leaves of G.
gracile has greater cytotoxic activity against various
cancer cell lines when compared to the apolar
extract.
A marked loss in cell viability is observed via MTT
assays after treatment of pancreatic, colon, and
prostate cancer cell lines with the ethanolic extract
of G. gracile leaves at concentrations between 25
and 125 µg/mL. The highest cytotoxic effect was
observed on pancreatic cancer Panc28 cells, and
prostate carcinoma androgen-dependent LNCaP
cells (figure 1A). Treatment with the apolar leaf
extract had a lesser effect on cell viability when
compared to the polar extract. However,
appreciable cytotoxic activity against colon cancer
HCT-116 cells is observed, followed by Panc 28,
androgen-independent prostate cancer PC-3, and
androgen-dependent LNCaP cells (figure 1B).
The polar extract derived from the inflorescences
of G. gracile inhibits cell viability on the cell lines
tested but not the apolar extract.
The inflorescence polar extract is most effective
at inhibiting cell viability of pancreatic cancer Panc
28 and colon cancer HCT-116 cells, with appreciable
activity on pancreatic MIA PaCa cancer cells at
concentrations between 25 and 125ug/mL (figure
1C). On the contrary, the apolar extract derived from
inflorescences has no effect on these cell lines, and
only has a modest effect on androgen-independent
PC-3 (figure 1D).
The polar and apolar extracts derived from
leaves and inflorescences of G. gracile have a
differential cytotoxic effect on the various cancer
cell lines tested
The polar extract from leaves effectively inhibits
cell viability in androgen-dependent LNCaP and
androgen-independent PC-3 prostate cancer cells,
pancreatic cancer MIA PaCa and colon
adenocarcinoma CaCo -2 cells (figure 1A). However,
the polar extract derived from inflorescences has a
minimal effect on these cell lines at the
concentrations shown (figure 1C). The apolar extract
derived from leaves display marked cytotoxic
activity against pancreatic Panc28, MIA PaCa, and
colon HCT-116 cancer cells (figure 1C). On the
contrary, the apolar extract from inflorescences has
no effect on these cell lines (figure 1D). Cell viability
is greatly diminished in colon cancer CaCo-2 cells by
the apolar extract from leaves, whilst the extract
PhOL Torrenegra-Guerrero, et al. 116 (pag 113-122)
http://pharmacologyonline.silae.it
ISSN: 1827-8620
from inflorescences may cause cell proliferation
(figures 1C and D).
The polar extracts from leaves and inflorescences
of G. gracile are highly effective against pancreatic
cancer cells Panc28.
The polar extracts are effective against Panc28
and MIA PaCa pancreatic cancer cells (figures 2A
and B) with the lowest observed EC50 of 20.23±1.185
µg/mL on Panc28 cells (figure 2C). The inflorescence
polar extract is also similarly effective on Panc28
cells with an EC50 of 27.58±1.063 µg/mL, but display a
lesser effect on MIA PaCa with an EC50 of 57.54± 1.06
µg/mL. The apolar extracts display low or no effect
on cell viability in these cells, except for the leaf
extract, which is effective against Panc 28 with an
EC50 of 57.88± 1.075 µg/mL (figure 2).
The viability of colon cancer HCT-116 but not
CaCo-2 cells is effectively diminished by the polar
and apolar extracts from leaves and the polar
extract from inflorescences of G. gracile.
Certain extracts from G. Gracile suppress cell
viability more effectively in HCT-116 cells than in
CaCo-2 (figure 3A and B). HCT-116 cells treated with
the polar extracts from leaves and inflorescences
and the apolar extract from leaves display EC50
values of 33.95±1.058, 39.46±1.0617, and
40.78±1.0405 µg/mL. While CaCo-2 values are ~50,
86.24±2.5609, and 65.8±1.0463 µg/mL for the same
extracts (figure 3C). The apolar extract from
inflorescences has no effect on either cell line
(figure 3).
Cell viability in both androgen-independent PC-3
and androgen dependent LnCaP prostate cancer
cells is effectively suppressed by the polar extract
derived from leaves of G. gracile.
The EC50 values of 28.84±1.1766 for PC-3 and
˂25.00 µg/mL for LnCaP cells suggest a highly
effective suppression of cell viability by the polar
extract derived from leaves. The apolar extract also
exerts a significant effect in PC-3 cells with an EC50
of 48.66±1.1069 µg/mL, and a much lesser effect on
LnCaP cells as indicated by an EC50 of 60.14±1.1715
µg/mL. The polar and apolar extracts derived from
inflorescences are less effective in these cells.
Discussion
The cytotoxic activity of four crude extracts from
leaves and inflorescences of Gnaphalium gracile
H.B.K. were studied against six human cancer cell
lines derived from pancreas (MIA PaCa, Panc 28),
colon (HCT-116, CaCo-2) and prostate (PC-3, LnCaP).
Our findings indicate that the differential cytotoxic
activity exerted by the extracts from leaves and
inflorescences are likely due to the various
compounds present in them as related to gene
expression profiles in these cell lines. This is
particularly evident by the significantly higher
activity displayed by the same extracts on colon
cancer HCT-116 as compared to CaCo-2 cells.
Additionally, the extracts suppressed cell viability in
all cells studied at various degrees of activity, except
for CaCo-2 cells, which were consistently least
affected.
This may be due to the fact that CaCo-2 is a better
differentiated cancer cell line with apicobasal
polarity similar to normal intestinal cells [15] and
that G. gracile is considered a non-toxic medicinal
plant, recommended to be taken in infusions.
Additionally, it has high concentrations of
flavonoids, among these, quercetin [13], a
recognized pro-apoptotic compound that targets
specifically and almost exclusively tumor cells, while
sparing normal cells [16, 17].
In our previous studies we have shown plant
derived compounds with preferential cytotoxic
activity towards cells categorized as highly
tumorigenic [10]. Aldehyde dehydrogenase (ALDH)
is a specific marker of subpopulations of cancer-
initiating cells in a tumor, and an indicator of
tumorigenic status in cancer cell lines [18-21]. The
effect G. gracile extracts on cell viability is observed
in cells categorized as highly tumorigenic as well as
those that are not, suggesting the presence of
compounds with varying targets. For example, the
polar leaf extract (EH polar) is most effective
against Panc28, LnCaP, PC-3, HCT-116, and MIA PaCa,
with half maximal effective concentrations between
(20.23±1.185 and 34.41±1.057µg/mL). Panc28, HCT-
116, and MIA PaCa express high levels of ALDH and
are categorized as highly tumorigenic [22-25], but
not LnCaP and PC-3 cells [26]. These half maximal
effective concentrations are the lowest obtained
among all extracts tested. Thus, the polar leaf
extract (EH) has the highest activity against the cell
lines tested. On the contrary, the apolar extract
from inflorescences (EF) has the least activity with
no effect on pancreatic or colon cancer cells,
suppressing cell viability only on PC-3 cells at a half
PhOL Torrenegra-Guerrero, et al. 117 (pag 113-122)
http://pharmacologyonline.silae.it
ISSN: 1827-8620
maximal effective concentration of 62.07±1.237
µg/mL. Indeed, this extract contains a high
concentration of low polarity substances such as
aromatic monoterpene compounds and fatty acids,
as well as low concentrations of flavonoids that may
be responsible for the observed weak activity on PC-
3 prostate cancer cells
The cytotoxic activity exerted by the G. gracile
crude extracts reported here is significantly higher
as compared to other reports of plants from the
genus Gnaphalium commonly advocated to have
antineoplastic properties. Extracts from the leaves,
stems, and inflorescences Gnaphalium spicatum are
reported to have IC50 values of >250µg/mL for most
cancer cells tested, with the most effective values
observed in the ethanolic extract obtained from the
roots with IC50 values ranging between 46 – 215
µg/mL for cells lines tested [27].
The polar extracts from leaves and
inflorescences, as well as the apolar extracts differ
greatly in activity towards the tumor cell lines
tested. This may be the result of the presence of
compounds at various concentrations, as well as to
differential distribution of compounds to certain
parts of the plant. Further research is needed to
isolate and determine the distribution and
concentration of these compounds in the plant, as
well as determine their specific targets and
mechanisms of action responsible for their
antineoplastic properties.
Acknowledgments
To the Universidad de Ciencias Aplicadas y
Ambientales U.D.C.A and East Tennessee State
University for financial support.
References
1. Rzedowski, G. C. de y J. Rzedowski. Flora
fanerogámica del Valle de México. 2a ed.
Instituto de Ecología y Comisión Nacional para el
Conocimiento y Uso de la Biodiversidad.
Pátzcuaro, Michoacán, México. 2001.
2. García-Barriga H. Flora Medicinal de Colombia
Nacional I, Editor. Bogotá, Colombia. 1975
3. Escarria S, Torrenegra R, Angarita B.
Colombian plants of the genus Gnaphalium.
Phytochemistry. 1977; 16: 1678.
4. Torrenegra RD. Quimica Y actividad biologica
de algunas plantas promisorias colombianas.
Revista Latinoamericana de Química. 2000; 28.
5. Torrenegra R, Escarria S, Achenbach H. Plantas
Colombianas del Genero Gnaphalium (II) Revista
Latinoamericana de Quimica. 1979; 10: 83-84.
6. Torrenegra R, Escarria S. Flavonoides de
Gnaphalium pellitum. Revista Latinoamericana de
Quimica. 1978; 8: 101.
7. Guerreiro E, Kavka J, Giordano O. 5,8-
dihydroxy-3,6,7-trimethoxyflavone from
Gnaphalium gaudichaudianum. Phytochemistry.
1982; 21: 2601-2602.
8. Bohlmann F, Ziesche J. Neue diterpene aus
Gnaphalium-arten. Phytochemistry. 1980; 19: 71-
74.
9. Torrenegra RD, Pedrozo J. Estudio
Quimiotaxonomico de los generos Gnaphalium y
Achyrocline Revista Asociacion Colombiana de
Ciencias Biologicas. 1984; 2: 39-44.
10. Torrenegra R, Pedrozo J, Rojas C, Carrizosa S.
Plantas Colombianas del genero Gnaphalium (IV)
G. rufescens y G. antennaroides Revista
Latinoamericana de Quimica. 1987; 18, 116-118.
11. Duran E, Ramsauer VP, Ballester M,
Torrenegra RD, Rodriguez OE, et al. Qualitative
analysis of sequence specific binding of flavones
to DNA using restriction endonuclease activity
assays. Biopolymers. 2013; 99: 530-537.
12. Thomas CM, Wood RC, 3rd, Wyatt JE,
Pendleton MH, Torrenegra RD, et al. Anti-
neoplastic activity of two flavone isomers
derived from Gnaphalium elegans and
Achyrocline bogotensis. PLoS One. 2012; 7:
e39806.
13. Torrenegra R, Ricardo A, Pedrozo J, Fuentes
O. Flavonoids from Gnaphalium gracile H.B.K.
Journal of Crude Drug Research. 1989; 27: 22.
14. Sclabas GM, Fujioka S, Schmidt C, Evans DB,
Chiao PJ. NF-kappaB in pancreatic cancer. Int J
Gastrointest Cancer. 2003; 33: 15-26.
15. Gilbert T, Rodriguez-Boulan E. Induction of
vacuolar apical compartments in the Caco-2
intestinal epithelial cell line. J Cell Sci. 1991; 100
(Pt 3): 451-458.
16. Park MH, Min do S. Quercetin-induced
downregulation of phospholipase D1 inhibits
proliferation and invasion in U87 glioma cells.
Biochem Biophys Res Commun. 2011; 412: 710-715.
PhOL Torrenegra-Guerrero, et al. 118 (pag 113-122)
http://pharmacologyonline.silae.it
ISSN: 1827-8620
17. Du G, Lin H, Wang M, Zhang S, Wu X, et al.
Quercetin greatly improved therapeutic index of
doxorubicin against 4T1 breast cancer by its
opposing effects on HIF-1alpha in tumor and
normal cells. Cancer Chemother Pharmacol. 2010;
65: 277-287.
18. Lee SH, Ryu JK, Lee KY, Woo SM, Park JK, et
al. Enhanced anti-tumor effect of combination
therapy with gemcitabine and apigenin in
pancreatic cancer. Cancer Lett. 2008; 259: 39-49.
19. Shin SY, Hyun J, Yu JR, Lim Y, Lee YH. 5-
Methoxyflavanone induces cell cycle arrest at the
G2/M phase, apoptosis and autophagy in HCT116
human colon cancer cells. Toxicol Appl
Pharmacol. 2011; 254: 288-298.
20. Erhart LM, Lankat-Buttgereit B, Schmidt H,
Wenzel U, Daniel H, et al. Flavone initiates a
hierarchical activation of the caspase-cascade in
colon cancer cells. Apoptosis. 2005; 10: 611-617.
21. Torres F, Quintana J, Estevez F. 5,7,3'-
trihydroxy-3,4'-dimethoxyflavone-induced cell
death in human leukemia cells is dependent on
caspases and activates the MAPK pathway. Mol
Carcinog 2010; 49: 464-475.
22. Neumeister V, Agarwal S, Bordeaux J, Camp
RL, Rimm DL. In situ identification of putative
cancer stem cells by multiplexing ALDH1, CD44,
and cytokeratin identifies breast cancer patients
with poor prognosis. Am J Pathol. 2010; 176: 2131-
2138.
23. Lin L, Liu Y, Li H, Li PK, Fuchs J, et al. Targeting
colon cancer stem cells using a new curcumin
analogue, GO-Y030. Br J Cancer. 2011; 105: 212-
220.
24. Visus C, Wang Y, Lozano-Leon A, Ferris RL,
Silver S, et al. Targeting ALDH(bright) human
carcinoma-initiating cells with ALDH1A1-specific
CD8(+) T cells. Clin Cancer Res. 2011; 17: 6174-
6184.
25. Schumacher M, Hautzinger A, Rossmann A,
Holzhauser S, Popovic D, et al. Potential role of P-
gp for flavone-induced diminished apoptosis and
increased adenoma size in the small intestine of
APC(min/+) mice. Cancer Invest. 2011; 29: 396-
404.
26. Karna P, Gundala SR, Gupta MV, Shamsi SA,
Pace RD, et al. Polyphenol-rich sweet potato
greens extract inhibits proliferation and induces
apoptosis in prostate cancer cells in vitro and in
vivo. Carcinogenesis. 2011; 32: 1872-1880.
27. Callacondo-Riva D, Quispe-Mauricio A, Lindo-
Gamarra S, Vaisberg AJ. Actividad citotoxica del
extracto etanolico de Gnaphalium spicatum
"keto keto" en cultivos de lineas celulares
tumorales humanas Rev Peru Med Exp Salud
Publica. 2008; 25: 380-385.
PhOL Torrenegra-Guerrero, et al. 119 (pag 113-122)
http://pharmacologyonline.silae.it
ISSN: 1827-8620
O
OH
O
OH
O
O
CH3
CH3
Quercetine
O
OH
OH
OH
O
OH
O
CH3
3 methoxy quercetine
O
OH
OH
OH
OH
OH
O
Gnaphaline
Figure 1. Molecular structures of Gnaphalin, 3-methoxy quercetine and quercetine.
Figure 2. Comparison of the effects of polar and apolar extracts derived from leaves and inflorescences on pancreatic, colon,
and prostate cancer cells. The effects of the polar and apolar extracts from leaves (1A and 1B) and inflorescences (1C and 1D)
on pancreatic (Mia Paca and Panc 28), colon (HCT 116 and Caco 2), and prostate (PC-3 and LNCaP) cancer cells were
determined by MTT assay and are represented as a percent of the control absorbance at a wavelength of 590 nm. All data
were collected at 24 h after treatment. Data shown are from representative experiments (n = 3). Values are expressed as
mean ± SE, * p˂0.05, significant difference between control and assayed concentrations for each extract.
PhOL Torrenegra-Guerrero, et al. 120 (pag 113-122)
http://pharmacologyonline.silae.it
ISSN: 1827-8620
Figure 3. The effects of polar and apolar extracts derived from leaves (EH) and inflorescences (EF) on pancreatic cancer cells.
Poorly differentiated MIA PaCa (figure 2A), and better differentiated Panc 28(figure 2B) pancreatic cancer cells were treated
with 25, 50 or 125 µg/ml of polar and apolar EH and EF extracts. The effects of the extracts on cell viability were determined
24 hours after treatment via MTT assay and are represented as a percent of the control absorbance at a wavelength 590nm.
Data shown are from representative experiments (n = 3). Values are expressed as mean ± SE, * p˂0.05, significant difference
between control and assayed concentrations for each extract. C. Half maximal effective concentration (EC50) ±SE for
treatment with polar and apolar extracts from leaves and. inflorescences on pancreatic carcinoma cells. The values were
estimated by non-linear regression analysis.
PhOL Torrenegra-Guerrero, et al. 121 (pag 113-122)
http://pharmacologyonline.silae.it
ISSN: 1827-8620
Figure 4. The effects of polar and apolar extracts derived from leaves (EH) and inflorescences (EF) on colon cancer cells.
Poorly differentiated HCT-116 (figure 3A), and better differentiated CaCo-2 (figure 3B) colon cancer cells were treated with
polar and apolar EH and EF extracts at concentrations of 25, 50 or 125 µg/ml. The effects of the extracts on cell viability were
determined by MTT assay 24 hours after treatment, and are represented as a percent of the control absorbance at a
wavelength 590nm. Data shown are from representative experiments (n = 3). Values are expressed as mean ± SE, * p˂0.05,
significant difference between control and assayed concentrations for each extract. C. Half maximal effective concentration
(EC50) ±SE for treatment with polar and apolar extracts from leaves and inflorescences on colon cancer cells. The values
were estimated by non-linear regression analysis.
PhOL Torrenegra-Guerrero, et al. 122 (pag 113-122)
http://pharmacologyonline.silae.it
ISSN: 1827-8620
Figure 5. The effects of polar and apolar extracts derived from leaves (EH) and inflorescences (EF) on prostate cancer cells.
Androgen-independent PC3 (figure 4A), and androgen-dependent LnCaP (figure 4B) prostate cancer cells were treated with
polar and apolar EH and EF extracts at concentrations of 25, 50 or 125 µg/ml. The effects of the extracts on cell viability were
determined by MTT assay 24 hours after treatment and are represented as a percent of the control absorbance at a
wavelength 590nm. Data shown are from representative experiments (n = 3). Values are expressed as mean ± SE, * p˂0.05,
significant difference between control and assayed extracts at specified concentrations. C. Half maximal effective
concentration (EC50) ±SE for treatment with polar and apolar extracts from leaves and inflorescences on colon cancer cells.
The values were estimated by non-linear regression analysis.
