Gibbilimbol analogues as antiparasitic agents - Synthesis and biological activity against Trypanosoma cruzi and Leishmania (L.) infantum

Article abstract of DOI:10.1016/j.bmcl.2016.01.040

The essential oils from leaves of Piper malacophyllum (Piperaceae) showed to be mainly composed by two alkenylphenol derivatives: gibbilimbols A and B. After isolation and structural characterization by NMR and MS data analysis, both compounds were evaluated against promastigote/amastigote forms of Leishmania (L.) infantum as well as trypomastigote/amastigote forms of Trypanosoma cruzi. The obtained results indicated that gibbilimbol B displayed potential against the tested parasites and low toxicity to mammalian cells, stimulating the preparation of several quite simple synthetic analogues in order to improve its activity and to explore the preliminary structure-activity relationships (SAR) data. Among the prepared derivatives, compound LINS03003 (n-octyl-4-hydroxybenzylamine) displayed the most potent IC₅₀ values of 5.5 and 1.8 μM against amastigotes of T. cruzi and L. (L.) infantum, respectively, indicating higher activity than the natural prototype. In addition, this compound showed remarkable selectivity index (SI) towards the intracellular forms of Leishmania (SI = 13.1) and T. cruzi (SI = 4.3). Therefore, this work indicated that preparation of synthetic compounds structurally based in the bioactive natural products could be an interesting source of novel and selective compounds against these protozoan parasites.

Full text of DOI:10.1016/j.bmcl.2016.01.040

Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Gibbilimbol analogues as antiparasitic agents—Synthesis and
biological activity against Trypanosoma cruzi and Leishmania (L.)
infantum
Marina T. Varela a, Roberto Z. Dias a, Ligia F. Martins b, Daiane D. Ferreira b, Andre G. Tempone b,
Anderson K. Ueno a, João Henrique G. Lago a, , João Paulo S. Fernandes a,
⇑
⇑
a Instituto de Ciências Ambientais, Químicas e Farmacêuticas, Universidade Federal de São Paulo, Rua São Nicolau 210, Diadema, SP 09913-030, Brazil
b Centro de Parasitologia e Micologia, Instituto Adolfo Lutz, Av. Dr. Arnaldo 355, São Paulo, SP 01246-902, Brazil
a r t i c l e i n f o
a b s t r a c t
Article history:
The essential oils from leaves of Piper malacophyllum (Piperaceae) showed to be mainly composed by two
alkenylphenol derivatives: gibbilimbols A and B. After isolation and structural characterization by NMR
and MS data analysis, both compounds were evaluated against promastigote/amastigote forms of
Leishmania (L.) infantum as well as trypomastigote/amastigote forms of Trypanosoma cruzi. The obtained
results indicated that gibbilimbol B displayed potential against the tested parasites and low toxicity to
mammalian cells, stimulating the preparation of several quite simple synthetic analogues in order to
improve its activity and to explore the preliminary structure–activity relationships (SAR) data. Among
the prepared derivatives, compound LINS03003 (n-octyl-4-hydroxybenzylamine) displayed the most
Received 15 December 2015
Revised 13 January 2016
Accepted 14 January 2016
Available online xxxx
Keywords:
Gibbilimbol A
Gibbilimbol B
Natural product derivatives
SAR
Leishmanicidal
Trypanocidal
potent IC50 values of 5.5 and 1.8 lM against amastigotes of T. cruzi and L. (L.) infantum, respectively, indi-
cating higher activity than the natural prototype. In addition, this compound showed remarkable selec-
tivity index (SI) towards the intracellular forms of Leishmania (SI = 13.1) and T. cruzi (SI = 4.3). Therefore,
this work indicated that preparation of synthetic compounds structurally based in the bioactive natural
products could be an interesting source of novel and selective compounds against these protozoan
parasites.
Ó 2016 Elsevier Ltd. All rights reserved.
Leishmaniasis and American trypanosomiasis (Chagas disease)
are tropical diseases caused by protozoan parasites belonging to
mastigotes. Our previous studies demonstrated that gibbilimbol B
affected the permeability of plasma membrane of L. (L.) infantum,
leading to cell death, a similar effect observed by the clinical used
drug amphotericin B. This action may be attributed to its amphi-
pathic characteristic that resembles membrane phospholipids,
where the phenolic hydroxyl can be directed to the aqueous layer
and the alkyl chain to the hydrophobic environment of membrane.
In continuation to our studies with P. malacophyllum, the essen-
tial oil from leaves showed to be composed by additional amounts
of gibbilimbol B as well as by its isomeric derivative gibbilimbol A
(Fig. 1). After several chromatographic steps,12 both compounds
were separated and their antiparasitic activities were individually
evaluated. As can be seen in Table 1, gibbilimbol A displayed weak
activity against amastigote forms of L. (L.) infantum and trypo-
Leishmania and Trypanosoma genus.1–4 Nowadays there is
a
reduced number of drugs to the treat these diseases, which include
highly toxic compounds such as pentamidine, amphotericin B, mil-
tefosine and nitroheterocyclic compounds (benznidazole and
nifurtimox), demonstrating the urgent necessity of novel and alter-
native treatments.5–7 Prospection of pharmacologically active
metabolites from plant species could be considered an interesting
approach to discovery of prototypes to be used in the development
of the new drugs to the treatment of parasitic diseases, mainly
those used in the ethnopharmacological point of view.8–10 In a pre-
vious work,11 our group reported the antiparasitic activity of gib-
bilimbol B, an alkylphenol derivative isolated from leaves of Piper
malacophyllum (Piperaceae) against promastigotes and amastig-
otes of Leishmania (L.) infantum, as well as Trypanosoma cruzi trypo-
mastigote forms of T. cruzi, with IC50 of 135.7 and 102.5 lM,
respectively. Otherwise, gibbilimbol B, an isomeric derivative of
gibbilimbol A (with double bond at C-40 instead of C-30) displayed
higher activity against the tested parasites and low toxicity to
mammalian cells. The result suggests the presence of unsaturation
close to aromatic ring leads to improved activity, and thus other
⇑
Corresponding authors. Tel.: +55 11 3385 4137.
E-mail addresses: joao.lago@unifesp.br (J.H.G. Lago), joao.fernandes@unifesp.br
(J.P.S. Fernandes).
http://dx.doi.org/10.1016/j.bmcl.2016.01.040
0960-894X/Ó 2016 Elsevier Ltd. All rights reserved.
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2
M. T. Varela et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
Considering the alkyl side chain nature of gibbilimbol deriva-
tives and the previous reported targeting ability to Leishmania
plasma membrane,11 interactions with specific membrane compo-
nents such as ergosterol could be plausible effect of these com-
pounds. Thus, eight analogues of gibbilimbols (LINS0300X, 1–8)
were designed (Fig. 1) to evaluate possible interactions by hydro-
gen-bonding and the role of the length of side chain (alkenyl or
alkyl) in the activity. Polar functional groups were inserted in this
moiety to provide additional interaction sites, so derivatives pos-
sessing carbonyl (1), hydroxyl (2) and amine (3) groups were
designed. As metabolic deactivation is also desirable in a future
drug to avoid excessive toxicity to mammalian cells,13 ester (4),
amide (5) and imine (6) derivatives were also proposed, since they
can be easily hydrolyzed in vivo. Moreover, lower homologues of
these compounds (7 and 8) were also synthesized and evaluated.
Compounds 1–8 were synthesized according to the Schemes 1
and 2, through classical methods which conducted to excellent
yields.14 Compound 1 was obtained through aldol condensation
of 4-hydroxybenzaldehyde and 2-nonanone under basic condi-
tions.15 This compound was then used to obtain 2 by reduction
of carbonyl group using sodium triacetoxyborohydride (STAB) pre-
pared in situ prior to reduction.16 Interestingly, only the product of
HO
HO
gibbilimbol A
O
gibbilimbol B
OH
HO
HO
HO
LINS03001
LINS03002
O
N
H
O
HO
LINS03004
LINS03003
O
N
H
N
HO
HO
LINS03005
O
LINS03006
N
H
O
HO
HO
LINS03008
LINS03007
Figure 1. Structures of gibbilimbols A/B and related analogues (1–8).
1,2-reduction was observed. Generally
a,b-unsaturated ketones
can be reduced to both 1,2- and 1,4-reduction products when
reacted with borohydride.17 However, the distribution of these
products is variable. It is well known STAB is a 1,2-selective agent
functional groups in this region should be explored. Therefore, this
work aimed the preparation of synthetic compounds structurally
related to gibbilimbol B and the evaluation of the antiparasitic
activity against L. (L.) infantum/T. cruzi, extracellular and intracellu-
lar forms and mammalian cytotoxicity, as well as to explore the
structure–activity relationships (SAR).
in the
a
,b-unsaturated aldehydes reduction. However it is not a
good reducing agent for
a
,b-unsaturated ketones.18 Possibly the
extended conjugation to the aromatic ring played the role to obtain
selectively the 1,2-reduction product 2.
Table 1
Antiparasitic activities of gibbilimbols A/B, and of the synthetic derivatives 1–8 against trypomastigote/amastigote forms of T. cruzi and promastigote/amastigote forms of L. (L.)
infantum
Compounds
IC50
lM (95% CI)
CC50
l
M
SIb
(95% CI)
T. cruzi
T. cruzi
L. (L.) infantum
L. (L.) infantum
NCTC cellsa
TCT
TCA
LIP
LIA
trypomastigotes
amastigotes
promastigotes
amastigotes
1
11.8
(10.1–13.1)
6.1
(5.5–6.7)
17.0
(14.0–20.8)
4.5
(3.7–5.4)
26.5
(22.6–31.2)
NA
22.5
(19.1–26.5)
NA
82.7
(74.3–92.2)
125.1
(90.3–174.4)
28.6
(25.3–32.6)
123.4
(114.0–133.0)
45.1
(38.8–52.3)
19.9
(17.5–22.6)
NA
NA
NA
82.8
(66.5–103.1)
49.5
(32.9–74.4)
23.5
(15.5–82.9)
130.3
(107.9–157.4)
45.4
(27.3–75.3)
99.2
(79.3–124.3)
167.8
(98.3–286.4)
>300
7.0
8.1
1.4
29.0
1.7
—
3.7
—
1.0
0.4
0.8
1.1
1.0
5.0
—
—
2
—
3
5.5
(4.6–6.5)
NA
1.8
(1.1–2.9)
NA
4.3
—
13.1
—
4
5
NA
NA
NA
NA
NA
NA
—
—
6
—
—
7
NA
NA
26.4
(20.4–33.3)
NA
—
6.4
—
—
8
31.6
(24.1–41.5)
NA
—
9.5
—
—
Gibbilimbol A
Gibbilimbol Bc
Miltefosine
Benznidazole
102.5
(74.5–122.7)
75.3
(62.3–90.9)
ND
NA
135.7
(92.1–163.5)
94.9
(73.2–123.2)
16.4
(15.4–17.4)
—
224.6
(201.3–256.9)
254.1
(217.9–296.3)
241.4
(206.9–281.6)
269.9
2.2
3.4
—
—
1.7
2.7
14.7
—
NA
100.4
(82.6–121.8)
16.7
(13.0–21.5)
—
—
2.5
14.5
—
ND
—
440.7
230.3
0.6
1.2
(406.1–478.3)
(176.2–301.1)
(414.9–532.1)
IC50: 50% inhibitory concentration.
NA: not active.
ND: not determined.
a
CC50: 50% cytotoxic concentration.
b
SI: selectivity index calculated from CC50/IC50 (TCT: T. cruzi trypomastigotes, TCA: T. cruzi amastigotes, LIP: L. (L.) infantum promastigotes, LIA: L. (L.) infantum
amastigotes).
c
Available from Oliveira et al.11
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M. T. Varela et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
3
O
OH
of 6.1 and 4.5 lM, respectively, it showed no activity against the
intracellular amastigotes. The lack of activity against intracellular
forms could be ascribed to low penetration in host cells, metabo-
lization inside macrophages generating inactive metabolites, but
also to enzymatic differences between trypomastigotes and
amastigotes,22 which may have resulted in low susceptibility of
the parasite to the gibbilimbol derivatives. When tested against
murine fibroblasts for mammalian cytotoxicity, compounds 1, 2
and 4 demonstrated selectivity when one considers the trypo-
mastigotes, with an index of 7.0, 8.1 and 29.0, respectively. The lat-
ter is the most selective compound in the series regarding the
trypomastigote form of T. cruzi. Compounds 1 and 3 demonstrated
selectivity index of 3.7 and 4.3 to intracellular amastigote form.
Based on the structures of most potent compounds, it is possible
to suggest that the presence of functional groups in the tail moiety
can significantly improve the activity, although a higher toxicity to
mammalian cells was also observed when compared to the natural
prototypes. However, derivatives 7 and 8 were not active in this
evolutionary form of the parasite (trypomastigotes), but have also
shown the lowest toxicity to mammalian cells among the designed
analogues. Trypomastigotes are clinical forms of the parasite abun-
dant in acute phase of the Chagas’ disease. Furthermore, drugs that
only target the replicating stages of the parasite may leave non-
replicating forms, such as trypomastigotes, capable of maintaining
infections long after the end of the treatment.23
(b)
5
5
HO
O
HO
(a)
1
2
N
(c)
N
H
(b)
n
n
HO
HO
HO
8: n = 3
3: n = 7
n = 3
6: n = 7
Scheme 1. Reagents and conditions of preparation of compounds 1–3, 6 and 8. (a)
2-nonanone, NaOH 40%, overnight; (b) STAB, AcOH, 4 h; (c) butylamine or
octylamine, THF, 60 °C, 1 h.
To obtain the imine derivatives 6 and 8, 4-hydroxybenzalde-
hyde and the corresponding amines were mixed together in
equimolar amounts, under ultrasonic irradiation.19 This reaction
led to quantitative conversion of the products and obviously excel-
lent yields. Starting from 6 the compound 3 was obtained through
reduction of the imine group using STAB.16 The lower homologue 8
was also obtained by the same way.
Finally, the esters 4 and 7 were obtained through classical Fis-
cher esterification using the 4-hydroxybenzoic acid and the corre-
sponding alcohol.20 In case of compound 4, 1-octanol was used in
excess in toluene, while 1-butanol was used as solvent in the syn-
thesis of 7. Otherwise, amide derivative 5 was obtained by the
reaction of 1-octylamine with the acyl chloride previously
obtained reacting the 4-hydroxybenzoic acid with thionyl
chloride.21
The antiprotozoal activity of gibbilimbol A and compounds 1–8
was evaluated in vitro against trypomastigote/amastigote forms of
T. cruzi as well as promastigote/amastigote forms of L. (L.) infantum
using the method from Oliveira et al.11 Benznidazole and miltefos-
ine were used as positive control against T. cruzi and L. (L.) infan-
tum, respectively. Cytotoxicity was determined in mammalian
NCTC clone 929 cells as previously described.11
Interestingly, the natural compound gibbilimbol B showed
weak activity against trypomastigotes and was not active against
T. cruzi amastigotes. The modifications proposed in this study
improved the effectiveness of this natural prototype, since the ana-
logues 1 and 3 presented not only activity, but also selectivity
towards the intracellular amastigotes of T. cruzi, which are respon-
sible for the chronic phase of the disease and a target stage consid-
ered for drug discovery studies.24 It is expected that a good
antitrypanosomal agent possess high potency (IC50 <10 lM)
against the amastigotes, together with adequate physicochemical
properties to achieve the intracellular environment.24 This can be
a possible explanation to the poor activity (and also to the low
cytotoxicity) of homologues 7 and 8, since they are less lipophilic
than the higher homologues and indeed may show poor penetra-
tion into the cells. Considering the effectiveness of compound 3
against both T. cruzi forms, further structural modifications may
contribute to the improvement of the biological selectivity.
Leishmania parasites were also susceptible to gibbilimbols and
their derivatives. Our data demonstrated that the isolated com-
pound gibbilimbol A was also active against the intracellular
Among the eight designed analogues, compounds 1–5 displayed
activity against the trypomastigotes of T. cruzi with IC50 values in a
range between 4 and 26
also presented activity, but showing
(IC50 = 102 M). The active compounds 1–5 showed 16–97-fold
l
M; the natural compound gibbilimbol A
a
reduced potential
l
more potency than the clinical used drug benznidazole against
the trypomastigotes (Y strain), suggesting the presence of func-
tional groups in the tail moiety can significantly influence the
activity. Although gibbilimbol B showed the weakest potency in
a previous study, it was 4-fold more active than benznidazole.
Among them, compounds 1 and 3 also demonstrated activity
against the intracellular amastigotes of T. cruzi, showing IC50 values
amastigotes (IC50 = 135.7 lM), but it was about 1.4-fold less potent
than the previous reported gibbilimbol B.11 Except for compound 7,
all derivatives demonstrated activity against the extracellular pro-
mastigotes of L. (L.) infantum, with IC50 values in a range between
of 22.5 and 5.5 lM, respectively. Although compounds 2 and 4
were the most potent against the trypomastigotes, with IC50 values
31 and 123 lM. The natural gibbilimbol A showed lack of activity
against the promastigotes. Compound 6 demonstrated the highest
activity against promastigotes, similarly to the standard drug mil-
tefosine. Among the designed derivatives, compound 3 demon-
strated potential against the intracellular amastigotes, with an
O
O
(a)
IC50 value of 1.8 lM, 52-fold higher potency against the intracellu-
n
lar amastigotes in comparison to natural prototype gibbilimbol B.
Furthermore, a high selectivity index (SI) value of 13 was found,
close to the standard drug miltefosine. Structural differences
between gibbilimbol B and derivative 3 suggested an improvement
on the antiparasitic effectiveness and conferred selectivity against
both Leishmania and T. cruzi intracellular amastigotes. Further
exploitation of 3 may contribute to improve the potency and
selectivity.
HO
O
7: n = 3
4: n = 7
OH
O
HO
N
H
(b)
HO
5
Our results suggest that the presence of a nitrogen atom in the
position C-20 of the side chain of phenolic derivatives significantly
improved the activity against L. (L.) infantum promastigotes, as
Scheme 2. Reagents and conditions to preparation of compounds 4, 5 and 7. (a) 1-
butanol or 1-octanol, H2SO4 (cat.), toluene, overnight; (b) SOCl2, triethylamine,
toluene, 60 °C, 2 h, then 1-octylamine, overnight.
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M. T. Varela et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
31.6, 31.4, 29.3, 22.5, 14.1; LREIMS m/z (rel. int.) 232 [M]+ (19), 133 (30), 121
(16), 120 (100), 107 (68); Gibbilimbol B: pale yellow oil; 1H NMR (300 MHz,
CDCl3, TMS, ppm) d 7.05 (d, 2H, J = 8.5 Hz), 6.75 (d, 2H, J = 8.5 Hz), 5.43 (m, 2H),
2.60 (t, 1H, J = 7.7 Hz), 2.27 (m, 2H), 1.98 (m, 2H), 1.27 (m, 8H), 0.89 (t, 3H,
J = 6.7 Hz); 13C NMR (75 MHz, CDCl3, TMS, ppm) d 153.4, 134.5, 131.2, 129.5,
129.3, 115.0, 35.2, 34.7, 32.6, 31.8, 29.5, 28.8, 22.6, 14.1; LREIMS m/z (rel. int.)
232 [M]+ (12), 204 (8), 133 (5), 120 (15), 107 (100).
observed to the compounds 3, 5, 6 and 8. Other substitution pat-
terns led to an activity close to the observed to natural gibbilimbols
A and B. Conversely, compounds presenting oxygen in C-20 posi-
tion, as well as other oxygenated functions directed to outside of
the tail (carbonyl or hydroxyl groups) led to poor activity. Consid-
ering the differences in the antiparasitic activity of natural proto-
types and the new derivatives, future studies towards the
discovery of mechanism of action may represent a tool to improve
the selectivity of these compounds.
In conclusion, although the toxicity of the gibbilimbol deriva-
tives was enhanced when compared to the natural scaffold (gibbil-
imbol B), our proposed modifications resulted in higher potency
against both parasites, also increasing the selectivity in the clini-
cally relevant form of these parasites (intracellular amastigotes).
In addition, the insertion of functional groups close to aromatic
ring led to more active compounds. Considering that gibbilimbol
B is a quite simple structure, further exploratory studies regarding
the mode of action of these compounds should be made, as well as
additional molecular modifications to enlarge the SAR data.
13. The Practice of Medicinal Chemistry; Wermuth, C. G., Ed.; Academic Press:
Oxford, 2008.
14. Yields and characterization data for the compounds 1: yellowish solid (40%); 1H
NMR (300 MHz, CDCl3, TMS, ppm) d 7.52 (d, 1H, J = 16.1 Hz), 7.46 (d, 2H,
J = 8.6 Hz), 6.87 (d, 2H, J = 8.6 Hz), 6.63 (d, 1H, J = 16.1 Hz), 5.84 (br s, 1H), 2.64
(t, 2H, J = 7.4 Hz), 1.55–1.75 (m, 2H), 1.21–1.42 (m, 8H), 0.88 (t, 3H, J = 6.8 Hz);
13C NMR (75 MHz, CDCl3, TMS, ppm) d 201.3, 158.0, 142.5, 130.2, 127.3, 124.0,
116.0, 40.9, 31.7, 29.3, 29.1, 24.6, 22.6, 14.1; HRMS calculated 246.1620; found
245.1560 [MꢁH+]; 2: yellowish oil (52%); 1H NMR (300 MHz, CDCl3, TMS, ppm)
d 7.27 (d, 2H, J = 8.0 Hz), 6.78 (d, 2H, J = 8.0 Hz), 6.50 (d, 1H, J = 15.6 Hz), 6.07
(dd, 1H, J = 15.6, 7.1 Hz), 4.88 (br s, 1H), 4.25 (q, 1H, J = 6.5 Hz), 1.50–1.72 (m,
4H), 1.16–1.48 (m, 8H), 0.88 (t, 3H, J = 6.7 Hz); 13C NMR (75 MHz, CDCl3, TMS,
ppm) d 159.3, 133.5, 130.7, 130.1, 130.0, 116.1, 71.8, 36.8, 31.9, 29.8, 29.5, 25.3,
22.7, 14.1; HRMS calculated 248.1776; found 247.1631 [MꢁH+]; 3: yellowish
oil (70%); 1H NMR (300 MHz, CDCl3, TMS, ppm) d 7.08 (d, 2H, J = 8.6 Hz), 6.60
(d, 2H, J = 8.6 Hz), 3.68 (s, 2H), 3.64 (br s, 2H), 2.67 (t, 2H, J = 7.3 Hz), 1.54
(quint., 2H, J = 7.1 Hz), 1.16–1.38 (m, 10H), 0.87 (t, 3H, J = 7.0 Hz); 13C NMR
(75 MHz, CDCl3, TMS, ppm) d 155.8, 130.6, 129.7, 115.8, 53.5, 49.5, 31.8, 29.6,
29.5, 29.2, 27.3, 22.6, 14.1; HRMS calculated 235.1936; found 236.2008 [M
+H+]; 4: colorless oil (82%); 1H NMR (300 MHz, CDCl3, TMS, ppm) d 7.96 (d, 2H,
J = 8.9 Hz), 6.87 (d, 2H, J = 8.9 Hz), 6.07 (br s, 1H), 4.28 (t, 2H, J = 6.7 Hz), 1.68–
1.83 (m, 2H), 1.20–1.50 (m, 10H), 0.88 (t, 3H, J = 7.0 Hz); 13C NMR (75 MHz,
CDCl3, TMS, ppm) d 166.8, 160.0, 131.9, 121.8, 115.2, 65.1, 31.8, 29.3, 29.2, 28.7,
26.1, 22.6, 14.1; HRMS calculated 250.1568; found 249.1512 [MꢁH+]; 5:
yellowish oil (71%); 1H NMR (300 MHz, CDCl3, TMS, ppm) d 7.60 (d, 2H,
J = 8.2 Hz), 6.87 (d, 2H, J = 8.2 Hz), 6.28 (t, 1H, J = 5.3 Hz), 3.42 (q, 2H, J = 6.6 Hz),
1.59 (quint., 2H, J = 7.2 Hz), 1.15–1.40 (m, 10H), 0.87 (t, 3H, J = 6.9 Hz); 13C
NMR (75 MHz, CDCl3, TMS, ppm) d 168.3, 160.2, 128.8, 125.6, 115.6, 40.3, 31.8,
29.3, 29.2, 27.0, 26.6, 22.6, 14.1; HRMS calculated 249.1728; found 248.1649
[MꢁH+]; 6: white solid (60%); 1H NMR (300 MHz, CDCl3, TMS, ppm) d 8.15 (s,
1H), 7.49 (d, 2H, J = 8.6 Hz), 6.67 (d, 2H, J = 8.8 Hz), 6.40 (br s, 1H), 3.57 (t, 2H,
J = 6.9 Hz), 1.68 (quint., 2H, J = 6.9 Hz), 1.30-1.52 (m, 10H), 0.86 (t, 3H,
J = 7.0 Hz); 13C NMR (75 MHz, CDCl3, TMS, ppm) d 162.4, 160.4, 130.3, 126.5,
116.0, 60.8, 31.8, 30.7, 29.3, 29.2, 27.2, 22.6, 14.1; HRMS calculated 233.1779;
found 232.1702 [MꢁH+]; 7: white solid (85%); 1H NMR (300 MHz, CDCl3, TMS,
ppm) d 7.95 (d, 2H, J = 8.1 Hz), 6.88 (d, 2H, J = 8.1 Hz), 6.14 (s, 1H), 4.30 (t, 2H,
J = 6.6 Hz), 1.80-1.69 (m, 2H), 1.47 (sext., 2H, J = 7.4 Hz), 0.97 (t, 3H, J = 7.4 Hz);
13C NMR (75 MHz, CDCl3, TMS, ppm) d 167.9, 159.9, 131.9, 122.8, 115.2, 64.8,
30.8, 19.3, 13.8; HRMS calculated 194.0942; found 193.0835 [MꢁH+]; 8:
yellowish oil (70%); 1H NMR (300 MHz, CDCl3, TMS, ppm) d 7.06 (d, 2H,
J = 8.1 Hz), 6.59 (d, 2H, J = 8.1 Hz), 5.48 (br s, 1H), 3.69 (s, 2H), 2.69 (t, 2H,
J = 7.4 Hz), 1.59–1.47 (m, 2H), 1.32 (sext., 2H, J = 7.2 Hz), 0.89 (t, 3H, J = 7.2 Hz);
13C NMR (75 MHz, CDCl3, TMS, ppm) d 156.3, 129.8, 129.7, 115.9, 53.4, 49.1,
31.5, 20.5, 13.9; HRMS calculated 179.1310; found 180.1408 [M+H+].
15. Ramachandra, M. S.; Subbaraju, G. V. J. Asian Nat. Prod. Res. 2006, 8, 683.
16. Tummatorn, J.; Dudley, G. B. Org. Lett. 2011, 13, 158.
Acknowledgements
The authors would like to thank CNPq (455411/2014-0,
470853/2012-3 and 471458/2012-0) and FAPESP (2015/11936-2,
2013/20479-9, 2012/18756-1) for providing financial support and
fellowship to M.T.V. (2014/16564-3). J.P.S.F., A.G.T. and J.H.G.L.
thanks to CNPq for the scientific research award.
References and notes
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12. Fresh leaves (ꢀ100 g) of P. malacophyllum were subjected to hydro-distillation
to afford 260 mg of crude essential oil. A portion of the leaf oil (200 mg) was
submitted to column chromatography over silica gel CH2Cl2 and CH2Cl2/MeOH
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Please cite this article in press as: Varela, M. T.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.01.040