Sunteți pe pagina 1din 11

Phytomedicine 21 (2014) 676681

Contents lists available at ScienceDirect

Phytomedicine
journal homepage:www.elsevier.de/phymed

Infections

Antiparasitic activity and effect of casearins isolated from Casearia sylvestris


on Leishmania and Trypanosoma cruzi plasma membrane
a b b,c
Diego Dinis Bou , Andr G. Tempone , rika G. Pinto ,
a a,
Joo Henrique G. Lago , Patricia Sartorelli
a b
Instituto de Cincias Ambientais, Qumicas e Farmacuticas, Universidade Federal de So Paulo, 09972-270 Diadema, SP, Brazil
Departamento de Parasitologia, Instituto Adolfo Lutz, 01246-000 So Paulo, SP, Brazil
c
Instituto de Medicina Tropical de So Paulo, Universidade de So Paulo, Av. Dr. Enas de Carvalho Aguiar, 470, 05403-000 So Paulo, SP, Brazil
article info abstract

Article history: Leishmaniasis and Chagas disease are infectious diseases caused by parasite Leishmania sp. and Trypanosoma cruzi,
Received 3 September 2013 respectively, and are included among the most neglected diseases in several underde-veloped and developing countries, with
Received in revised form 25 October 2013 an urgent demand for new drugs. Considering the antiparasitic potential of MeOH extract from leaves of Casearia sylvestris
Accepted 14 January 2014 Sw. (Salicaceae), a bioguided fractionation was conducted and afforded four active clerodane diterpenes (casearins A, B, G,
and J). The obtained results indicated a superior efficacy of tested casearins against trypomastigotes of T. cruzi, with IC 50
Keywords:
values ran-ging from 0.53 to 2.77 g/ml. Leishmania infantum promastigotes were also susceptible to casearins, with IC 50
Casearia sylvestris
Salicaceae values in a range between 4.45 and 9.48 g/ml. These substances were also evaluated for mam-malian cytotoxicity against
Clerodane diterpenes NCTC cells resulting in 50% cytotoxic concentrations (CC 50 ) ranging from 1.46 to 13.76 g/ml. Additionally, the action of
Leishmania casearins on parasite membranes was investigated using the fluorescent probe SYTOX Green. The obtained results
Trypanosoma cruzi demonstrated a strong interaction of casearins A and B to the plasma membrane of T. cruzi parasites, corroborating their
Casearins higher efficacy against these parasites. In contrast, the tested casearins induced no alteration in the permeability of plasma
membrane of Leishmania parasites, suggesting that biochemical differences between Leishmania and T. cruzi plasma
membrane might have contributed to the target effect of casearins on trypomastigotes. Thus, considering the importance of
studying novel and selective drug candidates against protozoans, casearins A, B, G, and J could be used as tools to future drug
design studies.

2014 Elsevier GmbH. All rights reserved.

Introduction countries (WHO 2010). The chemotherapy for these diseases is unsatisfactory
in terms of lack of effectiveness and also the unde-sirable side effects
Casearia sylvestris a plant popularly known as guac atonga, is associated with long term treatment with discovered drugs (Schmidt et al.
geographically distributed throughout Latin America (Lorenzi and Matos 2012). Medicinal plants have been used for the treatment of cutaneous
2002) and has been used in traditional medicine as anti-inflammatory, anti- leishmaniasis by rural peo-ple (Fournet et al. 1992; Franca et al. 1996) and
ulcer, anti-ophidian and anti-tumor (Ferreira et al. 2011). The chemical have attracted more interest from the scientific community. Several
composition of C. sylvestris has been char-acterized by clerodane type compounds isolated from Brazilian plants have been described with potent
diterpenes, known as casearins A-X which showed pronounced antitumor anti-trypanossomal and antileishmanial activities (Schmidt et al. 2012).
activity (Itokawa et al. 1990; Morita et al. 1991; Carvalho et al. 1998; Wang However, only eleven derivatives displayed comparable activity to standard
et al. 2009a; Santos et al. 2010). Clerodane diterpenes with different drugs used in therapy to treat these tropical diseases, as could be seen in Table
stereochemistry from casearins, named casearvestrins A-C and other 1.
clerodane diterpenes have also been described from leaves of C. sylvestris
(Oberlies et al. 2002; Santos et al. 2007; Wang et al. 2009b). Mesquita et al. (2005) evaluated extracts of thirteen medici-nal plants from
the Brazilian Cerrado biome for antileishmanial (Leishmania donovani) and
Protozoan diseases as Leishmaniasis and Chagas disease are main health antitrypanosomal activities and discov-ered that the leaves extract of C.
and socioeconomic problems in many developing sylvestris exhibited activity at 100 g/mL. However, no active compounds have
been isolated so far.


Corresponding author. Tel.: +55 11 3319 3300; fax: +55 11 4043 6428. E-mail address: Thus, the present work describes for the first time the
psartorelli@unifesp.br (P. Sartorelli).
antileish-manial and antitrypanosomal activities of casearins
0944-7113/$ see front matter 2014 Elsevier GmbH. All rights reserved. A, B, G, and J
http://dx.doi.org/10.1016/j.phymed.2014.01.004
D.D. Bou et al. / Phytomedicine 21 (2014) 676681 677

Table 1
Brazilian plants and isolated compounds which displayed antiparasitic activity (anti-trypanossomal and antileishmanial) and comparison of IC 50 values with those reported to positive controls.

Scientific name (family) Parasite form IC50 values Reference

Active compounds Positive controls


Aspidosperma ramiflorum L. amazonensis (promastigotes) Ramiflorine B (4.9 g/ml) Pentamidine (10.0 g/ml) Tanaka et al. (2007)
(Apocynaceae)
Drimys brasiliensis T. cruzi (trypomastigotes) Polygodial (2.03 g/ml) Benznidazole (38.3 g/ml) Corra et al. (2011)
(Winteraceae)
Nectandra megapotamica T. cruzi (trypomastigotes) Machilin G (0.78 g/ml) Gentian violet (31.0 g/ml) Silva Filho et al. (2004)
(Lauraceae) Galgravin (1.64 g/ml)
Calopiptin (4.49 g/ml)
Ganschisandrine (4.54 g/ml)
Peschiera australis L. amazonensis (amastigotes) Coronaridine (4.7 g/ml) Glucantime (6.6 g/ml) Delorenzi et al. (2001)
(Apocynaceae)
Piper regnelli (Piperaceae) L. amazonensis (amastigotes) Eupomatenoid-5 (5.0 g/ml) Anphotericin (0.23 g/ml) Vendrametto et al. (2010)
Piper solmsianum T. cruzi (trypomastigotes) 3,4,3 ,4 -Dimethylenedioxy- Gentian violet (31.0 g/ml) Martins et al. (2003)
(Piperaceae) 5,5 -dimethoxy-7,7 -
epoxylignan
(3.47 g/ml)
Tanacetum parthenium L. amazonensis (promastigotes) Parthenolide (0.37 g/ml) Tiuman et al. (2005)
(Asteraceae)
Virola surinamensis T. cruzi (trypomastigotes) Grandisin (1.5 g/ml) Gentian violet Lopes et al. (1998)
(Myristicaceae)

isolated from the MeOH extract from the leaves of C. sylvestris using caserins G (77 mg) and J (51 mg). Fraction A12 (300 mg) was also submitted
bioguided fractionation procedures. Additionally, the effect on the to CC Sephadex LH-20 to yield casearin B (12 mg). Finally, fractionation of
permeability of plasma membrane of L. infantum and T. cruzi was evaluated. A13 (526 mg) over Sephadex LH-20, followed by prep TLC (hexanes 7:
EtOAC 3) afforded casearin A (43 mg) (Fig. 1).

Material and methods Parasite maintenance

General experimental procedures L. (L.) infantum (MHOM/BR/1972/LD) was maintained in Golden


hamsters, up to approximately 6070 days post-infection. Pro-mastigotes were
1 13 maintained in M-199 medium supplemented with 10% calf serum and 0.25%
H NMR and C spectra were recorded, respectively, at 300 and 75

MHz in a Bruker INOVA spectrometer. CD3 OD (Aldrich) was used as the hemin at 24 C. Trypomastigotes of T. cruzi (Y strain) were maintained in
solvent and as the internal standard. Silica gel (Merck, 230400 mesh), and LLC-MK2 (ATCC CCL 7) cells using Roswell Park Memorial Institute
Sephadex LH-20 were used for column chro-matographic separation, while (RPMI) 1640 medium sup-plemented with 2% calf serum at 37

C in a 5%
silica gel 60 PF254 (Merck) was used for analytical and preparative thin layer
CO2 incubator (Tada et al. 1986).
chromatography.

Plant material
Mammalian cells
Leaves of C. sylvestris were collected from a single tree at the Atlantic
The mammalian cells (NCTC clone 929) and LLC-MK2 were
Forest area in So Paulo city, SP, Brazil (coordinates 23 5308.86S, 46
maintained in M-199 medium without phenol red and supple-mented with
4010.45O), in October, 2012. The identification was made by Dr. Oriana
10% fetal bovine serum at 37 C in a 5% CO2 humidified incubator.
Fvero (Universidade Presbiteriana Mackenzie-SP).

Extraction Evaluation of antiparasitic activity

Leaves of C. sylvestris (290 g), were dried and grounded, and then In order to determine the 50% inhibitory concentration (IC 50 ), the isolated
extracted using MeOH for 3 days, to afford 11.1 g of crude extract. The active casearins were dissolved in 100% MeOH, serially diluted in culture medium
MeOH extract was suspended in MeOH:H2 O 2:1. After partition using and incubated with parasites at con-centrations in the range between 0.1 and
150 g/ml. The maximal concentration of MeOH in plate wells was 0.5% and
hexanes, CH2 Cl2 and EtOAc, were obtained the three phases which were
internal controls were used to detect possible solvent toxicity to parasites.
evaluated to anti-trypanosomal and anti-Lesihmania activities.
Leishmania. The isolated casearins were incubated with pro-mastigotes of
6
L. (L.) infantum seeded at 1 10 /well in 96 well plates. After 48 h, the
Separation, purification and identification of chemical viability of promastigotes was measured by the mitochondrial activity using
constituents the MTT assay at 570 nm (Tada et al. 1986). Pentamidine was used as a
positive control.
The active hexane phase (6.4 g) was subjected to SiO 2 col-umn Trypanosoma cruzi. Trypomastigotes of T. cruzi (Y strain) obtained from
chromatography gel eluted with increasing amounts of EtOAc in hexane (9:1 LLC-MK2 cells were counted in a Neubauer hemo-cytometer and seeded at 1
to 1:9) to give 23 fractions (A1A23). The active fractions A11, A12 and A13 106 /well in 96-well microplates. Ater 24 h, the viability of trypomastigotes
were submitted to new chromato-graphic steps guided by the antiparasitic was measured by the mito-chondrial activity using the MTT assay at 570 nm
activity. Thus, fraction A11 (380 mg) was fractionated over Sephadex LH-20, (Lane et al. 1996). Benznidazole was used as the standard drug.
eluted with MeOH and followed by SiO2 prep TLC (hexanes 8: EtOAC 2) to
afford
678 D.D. Bou et al. / Phytomedicine 21 (2014) 676681
12 16

13
11

20 14
MeO
1 9
17 15
2 10 8
3 5 7

4 6
O
19 R2
18
O
R1 O O

casearin R1 R2
A CH3CO2 OH ()
G CH3CO2 H
J n-C3H7CO2 OH ()
B CH3CO2 CH3CO2 ()
Fig. 1. Structures of casearins isolated from C. sylvestris.
Determination of the cytotoxicity 13
C NMR spectra of these fractions
against mammalian cells
showed in general from 29 to 31
The in vitro selectivity index signals, similar to those reported for
casearins in the literature.
(CC50 /IC50 ) was determined using
13
NCTC cells, which were seeded at Characteristic signals in C NMR
4 spectra were observed at 57
6 10 /well in 96 well plates and
M-199 medium as described above. (CH3 ) indicating the presence of a
The mammalian cells were methoxy group in the structure, and
incubated with casearins and the
72 (CH) for the carbinolic carbon
standard drugs pentamidine and
(C2) in all isolated com-pounds.
benznidazole for 48 h at 37 C in a The signals at 75 (CH) were
5% CO2 humidified incubator. The assigned to carbons attached to
viability of cells was determined by hydroxyl groups (C6), except to
the MTT assay as described above casearin G, which displayed the
and the 50% cytotoxic methylenic carbon (C6) at 34. The
concentration (CC50 ) was double bond at C-3 and C-4 also
calculated. characteristic of casearins was
observed at 123 and 142
Spectrofluorimetric detection of respectively. The chemical shifts at
disruption of the parasites 53 (C-5) and 35 (C-10) were
plasma membrane similar to those described for
casearins and analogous
Late growth-phase (non- compounds. The signals of
stationary) promastigotes were methynic carbons at 95 (C-18) and
washed in PBS, seeded at 2 97 (C-19) indi-cate the existence of
6 the diacetalic ring system C, while
10 /well and incubated with 1 M
those at 170174 suggest the

SYTOX Green for 15 min at 24 presence of acetate and butanoate
C (Mangoni et al. 2005). Casearins groups as substituents.
were added, and the fluorescence
was measured every 20 min for 60
min total. The maximum
permeabilization was obtained with
0.1% Triton X-100. Fluorescence Determination of
intensity was determined using a the 50% inhibitory
fluorimetric microplate reader concentration and
(FilterMax F5 Multi-Mode mammalian
Microplate Reader-Molecular cytotoxicity
Devices) with excitation and emis-
sion wavelengths of 485 and 520 Casearins were incubated for 24
nm, respectively. The following h with T. cruzi trypomastigo-tes and
internal controls were used in the 48 h with promastigotes of L. (L.)
evaluation: (i) the background infantum. The viability was
fluorescence of casearins at the determined by the colorimetric
respective wavelengths, (ii) the assay of MTT. All tested sub-
possible interference of DMSO, stances killed 100% of parasites at
(iii) untreated promastigotes, and the highest tested concentration
(iv) medium without any cells. (150 g/ml); casearins A, B, G and J
Samples were tested in triplicate. showed IC50 values in a range
between 0.53 and 2.7 g/ml against
T. cruzi trypomastigotes (Table 2).
The most promising activity was
Results observed for casearin J, which
showed an IC50 value of 0.53 g/mL.
Isolation and identification of Leishmania promastig-otes were
casearins also susceptible to casearins
resulting in IC50 values ranging
from 4 to 9 g/ml (Table 2).
The chromatographic
Similarly to T. cruzi assay, the most
procedures carried out to active active substance against Leishmania
fractions A11, A12 and A13 was casearin J, which showed an
allowed the isolation of four pure IC50 value of 4.45 g/ml (Table 2).
substances. The Casearins were also tested
for cytotoxicity after
incubation with NCTC cells
for 48 h. The viability was
also determined using the
MTT assay. Casearins
D.D. Bou et al. / Phytomedicine 21 (2014) 676681 679

Table 2
Antileishmanial, antitrypanosomal and cytotoxicity effects of casearins isolated from C. sylvestris.
Casearins IC50 ( g/ml) promastigotes IC50 ( g/ml) trypomastigotes CC50 ( g/ml)
L. (L.) infantum T. cruzi Cell NCTC
95% CI 95% CI 95% CI
A 6.34* (4.0110.01) 1.37* (1.071.75) 2.62 (1.923.59)
B 9.48* (7.4312.09) 2.77* (2.632.93) 13.76 (9.1920.58)
J 4.45* (3.935.04) 0.53* (0.350.81) 1.46 (1.361.58)
G 5.93* (5.596.29) 1.57* (1.232.00) 7.54 (5.849.74)

95% CI = 95% confidence interval; Leishmania (promastigotes) pentamidine IC 50 0.22 g/ml; T. cruzi (trypomastigotes) benznidazole IC50 114.68 g/ml. *p < 0.05 (compared to standard drug).

showed CC50 values in a range between 1 and 13 g/ml (Table 2). It was also among the tested substances was significant (p < 0.001). Methanol at 0.5%
possible to note that casearin J was the most cytotoxic substance. was used as internal control and showed no alteration in permeability of
membranes. Casearins showed minimum alter-ation in the permeability of
Leishmania membrane when compare to fully (100%) permeabilized parasites
Plasma membrane permeabilization after 60 min, as follows: (i) casearin J = 6%; (ii) casearin G = 2.8%; (iii)
casearin B = 4.6% and (iv) casearin A = 4%. According to ANOVA test, the
The activity of casearins on parasites membrane of trypomasti-gotes and difference among the tested substances was significant (p < 0.001).
promastigotes was evaluated using the fluorescent probe SYTOX green (Fig.
2). The isolated casearins resulted in an intense and time-dependent increase
in fluorescence levels when incu-bated with T. cruzi parasites. When
Discussion
compared to fully permeabilized (100%) parasites, casearins showed the
following percentage of permeability alteration on T. cruzi membranes after
Clerodane diterpenes have been isolated from different plant species such
60 min incuba-tion: (i) casearin J = 29%; (ii) casearin G = 58%; (iii) casearin
as Baccharis tricuneata (Wagner et al. 1978), Zuelania guidonia (Khan et al.
B = 80% and (iv) casearin A = 87%. According to ANOVA test, the difference
1990), Laetia procera (Jullian et al. 2005) and Laetia corymbulosa (Beutler et
al. 2000). These derivatives are char-acteristic of the genus Casearia and have
been previously described from several species such as Casearia rupestris
(Vieira-Jnior et al. 2011), Casearia obliqua (Vieira-Jnior et al. 2009) and
Casearia nigrencens (Williams et al. 2007). However, characteristic diter-
penes from C. sylvestris are known as caseaerins/casearvestrins and display
promising biological activities, such as chemopreventive effect and anti-tumor
action (Prieto et al. 2013; Ferreira et al. 2010 ). The structural identification of
casearins A, B, G and J, isolated in this work was performed by comparing the
1 13
data obtained from the spectral H and C NMR with the literature data
(Itokawa et al. 1990; Morita et al. 1991; Carvalho et al. 1998).

Folk uses and scientific investigations have highlighted the importance of


C. sylvestris extracts and their relevant bioactive potential, as anti-ulcer, anti-
inflammatory, anti-ophidian and anti-tumor potentialities (Ferreira et al. 2011).
Previous studies reported the antileishmanial and antitrypanosomal activity of
crude extracts from Brazilian Cerrado plants, including C. sylvestris
(Mesquita et al. 2005), but no information about the activity of isolated com-
pounds have been described. Thus, this work demonstrates for the first time
the in vitro activity of casearins A, B, G and J, iso-lated from C. sylvestris
against T. cruzi and Leishmania parasites. Our data demonstrated that tested
casearins were most effec-tive against T. cruzi parasites in comparison to
standard drug benznidazole. Although casearin J showed the most promising
activity against T. cruzi, casearins B and G were the most selective substances,
with a selectivity index (CC50 mammalian cells/IC50 parasite) determined as
approximately 5. Considering the struc-tural differences among the isolated
casearins, we suggest that the substituent butanoate at C-18 present in casearin
J might have con-tributed to the higher efficacy among the four tested
diterpenoids. Despite no antitrypanosomal activity of casearins has previously
been described in the literature, a related clerodane diterpene iso-lated from C.
sylvestris showed a similar activity on T. cruzi, with minimal inhibitory
concentration (MIC) at 0.59 g/mL (Espindola et al. 2004).

Fig. 2. Evaluation of the plasma membrane permeability of (A) Leishmania and


(B) T. cruzi plasma membrane in the presence of casearins A, B, G, and J, assessed with the
vital dye SYTOX Green. Parasites were treated with Triton X-100 for 100% permeabilization. A Leishmania parasites were also susceptible to tested casearins, but to a
control group (untreated) was also included. The fluorescence was determined using a
fluorimetric microplate reader (FilterMax F5 Multi-Mode Microplate Reader) with excitation
lesser extent when compared to T. cruzi. According to the analysis of the IC 50
and emission wavelengths of 485 and 520 nm, respectively. values, tested casearin resulted in a lower efficacy than the standard drug
pentamidine. Similarly to the
680 D.D. Bou et al. / Phytomedicine 21 (2014) 676681
that casearins A and B showed Mangoni, M.L., Saugar, J.M., Dellisanti,
M., Barra, D., Simmaco, M., Rivas, L.,
activity observed against T. cruzi, the highest affinity to plasma Carvalho, P.R.F., Furlan, M., Young, M.C.M., 2005. Temporins, small antimicrobial
casearin J also demonstrated the membrane of T. cruzi, causing a Kingston, D.G.I., Bolzani, V.S., 1998. peptides with leishmanicidal activity. J.
Acetylated DNA damaging clerodane
highest efficacy among the tested strong permeabilization, which Biol. Chem. 280, 984990.
diterpenes from Casearia sylvestris. Phy- Marr, A.K., McGwire, B.S., McMaster,
substances against Leish-mania, may have contributed to parasite tochemistry 49, 16591662. W.R., 2012. Modes of action of
corroborating our above cited death. The presence of the Corra, D.S., Tempone, A.G., Reimo, J.Q., leishmanicidal antimicrobial peptides.
acetate substituent at C18 in Taniwaki, N.N., Romoff, P., Fvero,
suggestions about the structure O.A., Sartorelli, P., Mecchi, M.C., Lago,
Future Microbiol. 7, 10471059.
addition of oxy-genated C6 on Martins, R.C.C., Lago, J.H.G.,
activity relations. As a J.H.G., 2011. Anti-leishmanial and anti-
Albuquerque, S., Kato, M.J., 2003.
consequence of the elevated toxic- the structure of casearins A and B trypanosomal potential of polygodial
Trypanocidal tetrahy-drofuran lignans
may also have led to a superior isolated from stem barks of Drimys
ity to mammalian cells and weak from inflorescences of Piper
brasiliensis Miers (Winteraceae).
antileishmanial activity, casearins affinity to T. cruzi membrane. Parasitol. Res. 109, 231236.
solmsianum. Phytochemistry 64, 667
Although the tested casearins 670.
were tested only against the Delorenzi, J.C., Attias, M., Gattass, C.R.,
extracellular form of Leishmania. showed antileishmanial activity Andrade, M., Rezende, C., Pinto, A.C.,
Medina, J.M., Rodrigues, J.C., De Souza,
against promastigotes, no Hen-riques, A.T., Bou-Habib, D.C.,
To date, this is also the first report Saraiva, E.M.B., 2001. Antileishmanial
W., Atella, G.C., Barrabin, H., 2012.
describing the antileishmanial alteration in the permeability of activity of an indole alkaloid from
Tomatidine promotes the inhibition of
24-alkylated sterol biosynthesis and
activ-ity of casearins. the plasma membrane was Peschiera australis. Antimicrob. Agents
mitochon-drial dysfunction in
observed, suggesting a different Chemother. 45, 13491354.
Leishmania amazonensis
Targeting protozoan plasma lethal action on Leishmania para- Espindola, L.S., Vasconcelos Jnior, J.R., promastigotes. Parasitology 139,
sites, which might be exploited 12531265.
membranes has been considered a Mesquita, M.L., Marqui, P., de Paula,
promising approach to the study of for future studies. J.E., Mambu, L., Santana, J.M., 2004.
Mesquita, M.L., Desrivot, J., Bories, C.,
Trypanocidal activity of a new diterpene
novel drug candidates (Oliveira et from Casearia sylvestris var. lingua.
Fournet, A., Paula, J.E., Grellier, P.,
Espindola, L.S., 2005. Antileishmanial
al. 2012). When one considers the Planta Med. 70, 10931095.
and trypanocidal activity of Brazilian
differences between mam-malian Ferreira, P.M.P., Santos, A.G., Tininis, A.G.,
cerrado plants. Mem. Inst. Oswaldo
Costa, P.M., Cavalheiro, A.J., Bolzani,
plasma membrane and those of V.S., Moraes, M.O., Costa-Lotufo, L.V.,
Cruz 100 (7), 783787.
protozoan parasites, it is possible Morita, H., Nakayama, M., Kojima, H.,
Montenegro, R.C., Pessoa, C., 2010.
Casearin X exhibits cytotoxic effects in Takeya, K., Itokawa, H., Schenkel,
to envisage biochemical targets for Conclusion E.P., Motidome, M., 1991. Structures
leukemia cells triggered by apoptosis.
drugs. Sterols are the major Chem. Biol. Interact. 188, 497504. and cytotoxic activity relationship of
structural components of cell The obtained results Ferreira, P.M.P., Costa-Lotufo, L.V., Moraes, casearins, new clerodane diterpenes
from Casearia sylvestris Sw. Chem.
membranes and stabilize their indicated that the tested casearins M.O., Barros, F.W.A., Martins, A.M.A.,
Cavalheiro, A.J., Bolzani, V.S., Santos, Pharm. Bull. 39, 693697.
structure by interacting with the cause alteration in the A.G., Pessoa, C.O., 2011. Folk uses and
fatty acyl moieties of membrane permeability of plasma phar-macological properties of Casearia Oberlies, N.H., Burgess, J.P., Navarro,
phospholipids to affect membrane sylvestris: a medicinal review. An. Acad. H.A., Pinos, R.E., Fairchild, C.R.,
membrane of T. cruzi. Therefore, Peterson, R.W., Soejarto, D.D.,
fluidity. The sterol contents of Bras. Cienc. 83, 13731384.
considering the importance of Fournet, A., Angelo, A., Munoz, V., Roblot, Farnsworth, N.R., Douglas, K.A.,
microorganisms such as fungi and studying novel and selec-tive F., Hocquemiller, R., Cav, A., 1992. Wani, M.C., Wall, M.E., 2002. Novel
parasites of the Trypanosomati-dae Biolog-ical and chemical studies of Pera bioactive clerodane diterpenoids from
drug candidates against the leaves and twigs of Casearia
family differ from those of benensis, a Bolivian plant used in folk
protozoans, casearins could be medicine as a treatment of cutaneous sylvestris. J. Nat. Prod. 65, 9599.
mammalian cells because of the used as promising tools to future leishmaniasis. J. Ethnopharmacol. 37, Oliveira, A., Mesquita, J.T., Tempone,
predominant presence of drug design studies. 159164. A.G., Lago, J.H.G., Guimares, E.F.,
Kato, M.J., 2012. Leishmanicidal
ergosterol, episterol, and other 24- Franca, F., Lago, E.L., Marsden, P.D., 1996.
Plants used in the treatment of activity of an alkenylphenol from Piper
methyl sterols, which are malacophyllum is related to plasma
Acknowledgments leishmanial ulcers due to Leishmania
completely absent in the host cells (Viannia) braziliensis in an endemic area membrane disruption. Exp. Parasitol.
(Medina et al. 2012). Thus, the of Bahia, Brazil. Rev. Soc. Bras. Med. 132, 383387.
The authors thank FAPESP Trop. 29, 229232. Prieto, A.M., Santos, A.G., Oliveira, A.P.,
effect of casearins on the Cavalheiro, A.J., Silva, D.H.S.,
(2011/51739-0 and 2013/16320- Itokawa, H., Totsuka, N., Morita, H., Takeya,
permeability of T. cruzi and K., Litaka, Y., Schenkel, E.P., Motidome, Bolzani, V.S., Varanda, E.A., Soares,
4) and CNPq (470853/2012-3) C.P., 2013. Assessment of the
Leishmania plasma membranes M., 1990. New antitumor principles,
for financial support for the casearins A-F, from Casearia sylvestris chemopreventive effect of casearin B,
was evaluated using the fluo- a clerodane diterpene extracted from
develop-ment of this work. We Sw. (Flacourtiaceae). Chem. Pharm.
rescent probe SYTOX green. Our Bull. 38, 33843388. Casearia sylvestris (Salicaceae). Food
also thank CAPES and FAPESP
data clearly showed that casearins Jullian, V., Bonduelle, C., Valentin, A., Chem. Toxicol. 53, 153159.
fellowships to DDB and EGP, Santos, A.G., Perez, C.C., Tininis, A.G.,
have a higher affinity to T. cruzi Acebey, L., Duigou, A.-G., Prvost, M.-
respectively, and CNPq for the F., Sauvain, M., 2005. New clerodane Bolzani, V.S., Cavalheiro, A.J., 2007.
membrane than Leishmania, sug- Clerodane diterpenes from leaves of
scientific research award to diterpenoids from Laetia procera
gesting a time-dependent event (Poepp.) Eichler (Fla-courtiaceae), with Casearia sylvestris Swartz. Quim.
JHGL and AGT. The authors Nova 30, 11001103.
that might have contributed to cell antiplasmodial and antileishmanial
wish to thank Mr. Carlos R. activities. Bioorg. Med. Chem. Lett. 15, Santos, A.G., Ferreira, P.M.P., Vieira Jnior,
death. Membrane disruption G.M., Perez, C.C., Tininis, A.G., Silva,
Figueiredo for the collection of 50655070.
results in depletion of ionic Khan, M.R., Gray, A.I., Reed, D.R., Sadler, G.H., Bolzani, V.S., Costa-Lotufo,
Casearia sylvestris. L.V., Pessoa, C.O., Cavalheiro, A.J.,
gradients across the plasma I.H., Waterman, P.G., 1990. Diterpenes
from Zuelania guidonia. Phytochemistry 2010. Casearin X, its degradation
membrane, efflux of nutrients and product and other clerodane diterpenes
References 29, 16091614.
other cyto-plasmic components, Lane, J.E., Ribeiro-Rodrigues, R., Suarez, from leaves of Casearia sylvestris:
breakdown of parasite bioenergetic C.C., Bogitsh, B.J., Jones, M.M., Singh, evaluation of cytotoxicity against
Beutler, J.A., McCall, K.L., Herbert, K., normal and tumor human cells. Chem.
system and osmotic lysis, resulting Johnson, T., Shoemaker, R.H.,
P.K., Carter, C.E., 1996. In vitro
trypanocidal activity of tetraethylthiuram Biodivers. 7, 205215.
in cell death (Marr et al. 2012). Boyd, M.R., 2000. Cytotoxic
Oppo-sing to the observed clerodane diterpenes esters from
disulfide and sodium diethylamine-N- Schmidt, T.J., Khalid, S.A.,
carbodithioate on Trypanosoma cruzi.
antitrypanosomal effect, in which Laetia corymbulosa. Phytochemistry Am. J. Trop. Med. Hyg. 55, 263266. Romanha, A.J., Alves,
55, 233236.
casearin J presented the highest Lopes, N.P., Chicaro, P., Kato, M.J.,
Albuquerque, S., Yoshida, M., 1998.
T.M.A., Biavatti, M.W.,
antiparasitic activity, it is clear that Flavonoids and lignans from Virola Brun, R., Da Costa, F.B.,
this diter-pene induced a minimum surinamensis twigs and their in vitro
alteration of the plasma membrane activity against Trypanosoma cruzi. Castro, S.L., Ferreira, V.F.,
of T. cruzi when compared to other
Planta Med. 64, 667669. Lacerda, M.V.G., Lago,
Lorenzi, H., Matos, F.J.A., 2002. Plantas
tested casearins. Among these sub- medicinais do Brasil: Nativas e exticas. J.H.G., Leon, L.L., Lopes,
stances, our assays demonstrated Instituto Plantarum, Nova Odessa.
N.P., Neves, A.R.C.,
Niehues, M., Ogungbe, I.V.,
Pohlit, A.M., Scotti, M.T.,
Setzer,
D.D. Bou et al. / Phytomedicine 21 (2014) 676681 681
W.N., Soeiro, M.N.C., Steindel, M., Vieira-Jnior, G.M., Goncalves, T.O.,
Tempone, A.G., 2012. The potential of Ragasini, L.O., Ferreira, P.M.P., Pessoa,
sec-ondary metabolites from plants as C.O., Lotufo, L.V., Torres, R.B.,
drugs or leads against protozoan Boralle, N., Bolzani, V.S., Cavalheiro,
neglected diseases: Part I. Curr. Med. A.J., 2009. Cytotoxic clerodane
Chem. 19, 21282175. diterpenoids from Casearia obliqua. J.
Silva Filho, A.A., Albuquerque, S., Silva, Nat. Prod. 72, 18471850.
M.L.A., Eberlin, M.N., Tomazela, Vieira-Jnior, G.M., Dutra, L.A., Ferreira,
D.M., Bas-tos, J.K., 2004. P.M.P., Moraes, M.O., Costa Lotufo,
Tetrahydrofuran lignans from L.V., Pes-soa, C.O., Torres, R.B.,
Nectandra megapotamica with Boralle, N., Bolzani, V.S., Cavalheiro,
trypanocidal activity. J. Nat. Prod. 67, A.J., 2011. Cytotoxic clerodane
4245. diterpenes from Casearia rupestris. J.
Tada, H., Shiho, O., Kuroshima, K., Nat. Prod. 74, 776781.
Koyama, M., Tsukamoto, K., 1986. An Wagner, H., Seitz, R., Lotter, H., Herz, W.,
improved colorimetric assay for 1978. New furanoid ent-clerodanes
interleukin 2. J. Immunol. Methods 93, from Baccharis tricuneata. J. Org.
157165. Chem. 43, 33393345.
Tanaka, J.C.A., da Silva, C.C., Ferreira, Wang, W., Zhao, J., Wang, Y.H., Smillie,
I.C.P., Machado, G.M.C., Leonc, L.L., T.A., Li, X.C., Khan, I.A., 2009a.
de Oliveira, A.J.B., 2007. Diterpenoids from Casearia sylvestris.
Antileishmanial activity of indole Planta Med. 75, 14361441.
alkaloids from Aspidosperma ram- Wang, W., Zulfiqar, A., Li, X.C., Smillie,
iflorum. Phytomedicine 14, 377380. T.A., Guo, D.A., Khan, I.A., 2009b.
Tiuman, T.S., Ueda-Nakamura, T., Cortez, New clero-dane diterpenoids from
D.A.G., Dias Filho, B.P., Daz, J.A.M., Casearia sylvestris. Fitoterapia 80, 404
Souza, W., Nakamura, C.V., 2005. 407.
Antileishmanial activity of WHO, 2010. Working to Overcome the
parthenolide, a sesquiterpene lactone Global Impact of Neglected Tropical
isolated from Tanacetum parthenium. Diseases: First WHO Report on
Antimicrob. Agents Chemother. 49, Neglected Tropical Diseases. WHO,
176182. Geneva, pp. 184.
Williams, R.B., Norris, A., Miller, J.S.,
Vendrametto, M.C., dos Santos, A.O., Birkinshaw, C., Ratovoson, F.,
Nakamura, C.V., Dias Filho, B.P., Andriantsifer-ana, R., Rasamison, V.E.,
Cortez, D.A.G., Ueda-Nakamura, T., Kingston, D.G.I., 2007. Cytotoxic
2010. Evaluation of antileishmanial clerodane diterpenoids and their
activity of eupomatenoid-5, a hydrolysis products from Casearia
compound isolated from leaves of Piper nigrescens from the rainforest of
regnellii var. pallescens. Parasitol. Int. Madagascar. J. Nat. Prod. 70, 206209.
59, 154158.

S-ar putea să vă placă și