Synthesis, characterization and in vitro anticancer activity of novel 8,4’-oxyneolignan analogues

Article abstract of DOI:10.21577/0103-5053.20170075

Neolignans are a class of natural products with a wide range of biological effects. These substances are of great synthetic and biological interest, especially in searching for novel anticancer agents. In this paper, we report the synthesis of a new subclass of 8,4’-oxyneolignan analogues (Β-ketoethers and Β-ketoesters) and their cell viability assay on twenty four different cancer cells, among leukemias and carcinomas. Three compounds inhibited the growth of most human cancer cells. 2-Oxo-2-phenylethyl(2E)-3-[4-(2-oxo-2-phenylethoxy) phenyl]prop-2-enoate showed an antiproliferative activity superior to doxorubicin for U-87, U-138 MG and H1299 cell types and (E)-2-oxo-2-phenylethyl 3-(3-methoxy-4-(2-oxo-2-phenylethoxy)phenyl)acrylate was found to be very selective, demonstrating a growth inhibition of 92.0% against KG-1 cells. Furthermore, 1-oxo-1-phenylpropan-2-yl cinnamate exhibited significant inhibition activity in a range of 52.2 to 91.2% against twelve kinds of leukemia cell lines, revealing excellent results and very comparable to the reference drug.

Full text of DOI:10.21577/0103-5053.20170075

http://dx.doi.org/10.21577/0103-5053.20170075
J. Braz. Chem. Soc., Vol. 28, No. 11, 2229-2243, 2017.
Printed in Brazil - ©2017 Sociedade Brasileira de Química
0103 - 5053 $6.00+0.00
Article
Synthesis, Characterization and in vitro Anticancer Activity of Novel
8,4’-Oxyneolignan Analogues
Gisele C. Souza,a Gilberto C. Franchi Jr.,b Alexandre E. Nowill,b Lourivaldo S. Santos,c
Cláudio N. Alves,c Lauro E. S. Baratad and Carlos K. Z. Andrade*,a
aLaboratório de Química Metodológica e Orgânica Sintética, Instituto de Química,
Universidade de Brasília, 70910-900 Brasília-DF, Brazil
bCentro Integrado de Pesquisas Oncohematológicas da Infância,
Universidade Estadual de Campinas, 13083-872 Campinas-SP, Brazil
cInstituto de Ciências Exatas e Naturais, Universidade Federal do Pará, 66075-110 Belém-PA, Brazil
dLaboratório de Síntese Orgânica, Universidade Estadual de Campinas, 13083-872 Campinas-SP, Brazil
Neolignans are a class of natural products with a wide range of biological effects. These
substances are of great synthetic and biological interest, especially in searching for novel anticancer
agents. In this paper, we report the synthesis of a new subclass of 8,4’-oxyneolignan analogues
(β-ketoethers and β-ketoesters) and their cell viability assay on twenty four different cancer cells,
among leukemias and carcinomas. Three compounds inhibited the growth of most human cancer
cells. 2-Oxo-2-phenylethyl(2E)-3-[4-(2-oxo-2-phenylethoxy) phenyl]prop-2-enoate showed an
antiproliferative activity superior to doxorubicin for U-87, U-138 MG and H1299 cell types and
(E)-2-oxo-2-phenylethyl 3-(3-methoxy-4-(2-oxo-2-phenylethoxy)phenyl)acrylate was found to
be very selective, demonstrating a growth inhibition of 92.0% against KG-1 cells. Furthermore,
1-oxo-1-phenylpropan-2-yl cinnamate exhibited significant inhibition activity in a range of 52.2 to
91.2% against twelve kinds of leukemia cell lines, revealing excellent results and very comparable
to the reference drug.
Keywords: neolignans, antiproliferative activity, MTT assay, β-ketoester, β-ketoether
Introduction
coupling of allyl and/or propenyl phenols and are normally
found in plants of the Myristicacea family.6-8 Neolignans
have a wide range of biological effects such as antioxidant,9-14
antibacterial,15-17 anti-inflammatory,12,18-21 antifungal,22,23
anti-leishmanial,24-28 anti-trypanosomastid,27,28 and
anticancer activity,13,14,29,30 among others.31-34 Therefore, this
type of natural product is of great synthetic and biological
interest, especially in searching for novel anticancer
agents. In this paper, we report the synthesis of new
8,4’-oxyneolignan analogues and the cell viability assays
for different neoplasms, among leukemias and carcinomas.
Cancer is a generic term for a large group of diseases
that can affect any part of the body and remains a leading
cause of death worldwide. It is considered a public health
problem according to the World Health Organization, and
many efforts have been made towards its prevention and
cure. Treatment usually involves a series of interventions,
and approximately 90% of tumors can be treated with
antiblastic drugs,1,2 which makes chemotherapy the most
used treatment. However, these drugs are non-selective
and are toxic to healthy tissues, especially those of rapid
cell proliferation.2-5 As a consequence, there is an urgent
need for novel and effective drugs that act against cancer.
Several substances have been thoroughly studied for
their biological activities and a very promising class are the
neolignans, which are substances derived from the oxidative
Experimental
Chemistry
All reagents were purchased from commercial suppliers
and were used as received, unless otherwise specified.
Reactions were monitored by thin layer chromatography
*e-mail: ckleber@unb.br

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Synthesis, Characterization and in vitro Anticancer Activity of Novel 8,4’-Oxyneolignan Analogues
J. Braz. Chem. Soc.
(TLC) using aluminum plates from Merck (silica gel
60 F254). Melting points (mp) were determined using a
PFM II apparatus (model 382). Proton (1H) and carbon
(13C) nuclear magnetic resonance (NMR) spectra were
recorded on the following spectrometers: BrukerAC 250/P,
Varian Mercury Plus 300 MHz, BrukerAvance 600 MHz or
Varian Inova 500 MHz. Chemical shifts are reported in ppm
(d) with values relative to TMS used as internal standard.
High-resolution mass spectra (HRMS) were recorded
on a VG AutoSpec High Resolution Mass Spectrometer
(Micromass Company) or on a triple TOF 5600+ High
Resolution Mass Spectrometer (AB Sciex) with internal
calibration and direct solution (1 ppm in methanol).
(5.07 mmol) of 2,4-dichlorophenol, 1.00 g (5.02 mmol) of
phenacyl bromide and 1.25 g (9.04 mmol) of K2CO3 and
purified by recrystallization from acetone/hexane (42%
yield). The product obtained was a light brown crystalline
solid (mp 74-75 °C). 1H NMR (600 MHz, CDCl3) d 8.00
(d, J 7.2 Hz, 2H, Ar-H), 7.63 (t, J 7.2 Hz, 1H, Ar-H), 7.50
(t, J 7.2 Hz, 2H, Ar-H), 7.38 (d, J 3.0 Hz, 1H), 7.13 (dd, J1
9.0 Hz, J2 3.0 Hz, 1H, Ar-H), 6.78 (d, J 9.0 Hz, 1H, Ar-H),
5.34 (s, 2H, CH2); 13C NMR (150 MHz, CDCl3) d 193.7,
152.7, 134.3, 134.3, 130.4, 129.0, 128.3, 127.7, 127.0,
124.3, 115.1, 72.1. HREIMS m/z 302.9949 [M + Na]+
(calcd. for C14H10Cl2NaO2+, 302.9951).
2-(4-Nitro-phenoxy)-1-phenyl-propan-1-one (3c)
General procedure for the synthesis of 8,4’-oxyneolignan
Obtained according to the general procedure from
0.55 g (3.93 mmol) of 4-nitrophenol, 0.82 g (3.85 mmol)
of 2-bromopropiophenone and 0.96 g (6.93 mmol) of
K2CO3 and purified by recrystallization from methanol
(91% yield). The product obtained was a white crystalline
solid (mp 78-80 °C).1H NMR (600 MHz, CDCl3) d 8.14 (d,
J 9.0 Hz, 2H, Ar-H), 8.04 (d, J 7.8 Hz, 2H, Ar-H), 7.63 (t,
J 7.8 Hz, 1H, Ar-H), 7.51 (t, J 7.8 Hz, 2H, Ar-H), 6.90 (d,
J 9.0 Hz, 2H, Ar-H), 5.63 (q, J 6.6 Hz, 1H, CH), 1.78 (d,
J 6.6 Hz, 3H, CH3); 13C NMR (150 MHz, CDCl3) d 197.2,
162.4, 142.0, 134.2, 133.7, 129.0, 128.7, 126.0, 115.0,
76.9, 18.8. HREIMS m/z 294.0737 [M + Na]+ (calcd. for
C15H13NNaO4+, 294.0737).
analogues: β-ketoesters and β-ketoethers
A solution of 1.02 equivalent of phenols (or
0.51 equivalent of cinnamic acid derivatives) and
1.80 equivalent of anhydrous K2CO3 in anhydrous butanone
(4.5 mL of solvent per mmol of phenol or cinnamic acid
derivative) was stirred for 15 min at room temperature.After
this period, a solution of 1.00 equivalent of α-bromoketone
in anhydrous butanone (1.5 mL of solvent per mmol
of ketone) was added dropwise and the mixture was
stirred under reflux for 24 h. The solution was cooled
to room temperature, filtered, and the residue washed
with CHCl3. The solution was concentrated in vacuum
(to remove butanone), diluted with H2O, and extracted
with CHCl3 (3×). The organic extracts were combined,
washed with water, 5% NaOH solution, brine, dried over
Na2SO4 and then filtered and concentrated in vacuum. The
reaction products were purified by crystallization or column
chromatography.
2-Pentachlorophenyloxy-1-phenyl-ethanone (3d)
Obtained according to the general procedure from 0.68 g
(2.56 mmol) of pentachlorophenol, 0.50 g (2.51 mmol) of
phenacyl bromide and 0.62 g (4.52 mmol) of K2CO3 and
purified by recrystallization from methanol/CH2Cl2 (100%
yield). The product obtained was a white crystalline solid
(mp 125-127 °C). 1H NMR (600 MHz, CDCl3) d 7.97 (d,
J 7.2 Hz, 2H, Ar-H), 7.63 (t, J 7.2 Hz, 1H, Ar-H), 7.51 (t,
J 7.2 Hz, 2H,Ar-H), 5.30 (s, 2H, CH2); 13C NMR (150 MHz,
CDCl3) d 192.1, 151.2, 134.3, 134.2, 132.2, 130.2, 129.1,
128.3, 128.2, 74.8. HREIMS m/z 406.8745 [M + Na]+
(calcd. for C14H7Cl5NaO2+, 406.8752).
2-(4-Nitro-phenoxy)-1-phenyl-ethanone (3a)
Obtained according to the general procedure from
1.50 g (10.76 mmol) of 4-nitrophenol, 2.10 g (10.55 mmol)
of phenacyl bromide, 2.62 g (19.00 mmol) of K2CO3 and
purified by recrystallization from acetone/hexane (84%
yield). The product obtained was an orange crystalline solid
(mp 148-150 °C). 1H NMR (600 MHz, CDCl3) d 8.20 (d,
J 9.0 Hz, 2H, Ar-H), 7.99 (d, J 7.2 Hz, 2H, Ar-H), 7.66 (t,
J 7.2 Hz, 1H, Ar-H), 7.54 (t, J 9.0 Hz, 2H, Ar-H), 6.99 (d,
J 9.0 Hz, 2H,Ar-H), 5.43 (s, 2H, CH2); 13C NMR (150 MHz,
CDCl3) d 192.8, 163.0, 142.1, 134.4, 134.0, 129.1, 128.0,
125.9, 114.8, 70.6. HREIMS m/z 280.0581 [M + Na]+
(calcd. for C14H11NNaO4+, 280.0580).
1-Phenyl-2-m-tolyloxy-ethanone (3e)
Obtained according to the general procedure from 0.28 g
(2.56 mmol) of m-cresol, 0.50 g (2.51 mmol) of phenacyl
bromide and 0.62 g (4.52 mmol) of K2CO3 and purified by
recrystallization from methanol (69% yield). The product
obtained was a yellow crystalline solid (mp 70-72 °C).
1H NMR (600 MHz, CDCl3) d 8.01 (d, J 7.8 Hz, 2H, Ar-H),
7.61 (t, J 7.8 Hz, 1H,Ar-H), 7.50 (t, J 7.8 Hz, 2H,Ar-H), 7.16
(t, J 7.8 Hz, 1H,Ar-H), 6.80 (d, J 7.8 Hz, 1H,Ar-H), 6.78 (s,
1H,Ar-H), 6.74 (dd, J1 7.8 Hz, J2 1.8 Hz, 1H,Ar-H), 5.24 (s,
2-(3,5-Dichlorophenoxy)-1-phenylethanone (3b)
Obtained according to the general procedure from 0.83 g

Vol. 28, No. 11, 2017
Souza et al.
2231
2H, CH2), 2.32 (s, 3H, CH3); 13C NMR (150 MHz, CDCl3) d
194.7, 158.1, 139.7, 134.7, 133.8, 129.3, 128.8, 128.2, 122.5,
115.7, 111.6, 70.8, 21.5. HREIMS m/z 249.0891 [M + Na]+
(calcd. for C15H14NaO2+, 249.0886).
1-(4-Bromophenyl)-2-(3,5-dichlorophenoxy)ethanone (3i)
Obtained according to the general procedure from 0.30 g
(1.84 mmol) of 2,4-dichlorophenol, 0.50 g (1.80 mmol) of
2,4’-dibromoacetophenone and 0.45 g (3.24 mmol) of
K2CO3 and purified by recrystallization from methanol/
CH2Cl2 (39% yield). The product obtained was a white
crystalline solid (mp 99-101 °C). 1H NMR (600 MHz,
CDCl3) d 7.88 (d, J 8.4 Hz, 2H, Ar-H), 7.65 (d, J 9.0 Hz,
2H,Ar-H), 7.39 (d, J 2.4 Hz, 1H,Ar-H), 7.14 (dd, J1 9.0 Hz,
J2 2.4 Hz, 1H, Ar-H), 6.78 (d, J 9.0 Hz, 1H, Ar-H), 5.25
(s, 2H, CH2); 13C NMR (150 MHz, CDCl3) d 193.1, 152.4,
132.9, 132.2, 130.4, 129.8, 129.5, 127.6, 127.2, 124.2,
114.9, 72.1. HREIMS m/z 380.9042 [M + Na]+ (calcd. for
C14H9BrCl2NaO2+, 380.9056).
2-(3,5-Dichlorophenoxy)-1-phenylpropan-1-one (3f)
Obtained according to the general procedure from 0.39 g
(2.40 mmol) of 2,4-dichlorophenol, 0.50 g (2.35 mmol) of
2-bromopropiophenone and 0.58 g (4.23 mmol) of K2CO3
and purified by column chromatography in hexane/
acetate (80:20) (54% yield). The product was obtained
as a colorless oil. 1H NMR (600 MHz, CDCl3) d 8.08 (d,
J 7.2 Hz, 2H, Ar-H), 7.59 (t, J 7.2 Hz, 1H, Ar-H), 7.47 (t,
J 7.2 Hz, 2H, Ar-H), 7.35 (d, J 2.6 Hz, 1H, Ar-H), 7.06
(dd, J1 8.8 Hz, J2 2.6 Hz, 1H, Ar-H), 6.72 (d, J 8.8 Hz, 1H,
Ar-H), 5.40 (q, J 6.8 Hz, 1H, CH), 1.78 (d, J 8.8 Hz, 3H,
CH3); 13C NMR (150 MHz, CDCl3) d 194.7, 158.1, 139.7,
134.7, 133.8, 129.3, 128.8, 128.2, 122.5, 115.7, 111.61,
70.8, 21.5. HREIMS m/z 317.0115 [M + Na]+ (calcd. for
C15H12Cl2NaO2+, 317.0107).
1-(4-Bromophenyl)-2-(m-tolyloxy)ethanone (3j)
Obtained according to the general procedure from
0.10 g (0.92 mmol) of m-cresol, 0.25 g (0.90 mmol) of
2,4’-dibromoacetophenone and 0.23 g (1.62 mmol) of
K2CO3 and purified by recrystallization from methanol
(69% yield). The product obtained was a light brown
crystalline solid (mp 91-93 °C).1H NMR (600 MHz, CDCl3)
d 7.88 (d, J 8.4 Hz, 2H,Ar-H), 7.63 (d, J 8.4 Hz, 2H,Ar-H),
7.16 (t, J 8.0 Hz, 1H, Ar-H), 6.81 (d, J 8.0 Hz, 1H, Ar-H),
6.76 (s, 1H, Ar-H), 6.72 (d, J 8.0 Hz, 1H, Ar-H), 5.17 (s,
2H, CH2), 2.32 (s, 3H, CH3); 13C NMR (150 MHz, CDCl3)
d 194.1, 157.9, 139.8, 133.4, 132.1, 129.8, 129.3, 129.1,
122.7, 115.6, 111.5, 70.9, 21.5. HREIMS m/z 326.9997
[M + Na]+ (calcd. for C15H13BrNaO2+, 326.9992).
2-Pentachlorophenyloxy-1-phenyl-propan-1-one (3g)
Obtained according to the general procedure from 0.64 g
(2.40 mmol) of pentachlorophenol, 0.50 g (2.35 mmol) of
2-bromopropiophenone and 0.58 g (4.23 mmol) of K2CO3
and purified by recrystallization from methanol (66%
yield). The product obtained was a light brown crystalline
solid (mp 111-112 °C). 1H NMR (600 MHz, CDCl3) d 8.04
(d, J 7.2 Hz, 2H, Ar-H), 7.61 (t, J 7.2 Hz, 1H, Ar-H), 7.50
(t, J 7.2 Hz, 2H,Ar-H), 5.70 (q, J 7.0 Hz, 1H, CH), 1.68 (d,
J 7.0 Hz, 3H, CH3); 13C NMR (150 MHz, CDCl3) d 196.3,
150.7, 134.5, 133.9, 132.2, 129.7, 129.1, 128.9, 128.4,
82.1, 18.8. HREIMS m/z 420.8903 [M + Na]+ (calcd. for
C15H9Cl5NaO2+, 420.8908).
2-Oxo-2-phenylethyl(2E)-3-[4-(2-oxo-2-phenylethoxy)
phenyl]prop-2-enoate (4a)
Obtained according to the general procedure from
0.21 g (1.28 mmol) of p-hydroxycinnamic acid, 0.50 g
(2.51 mmol) of phenacyl bromide and 0.62 g (4.52 mmol) of
K2CO3 and purified by recrystallization from acetone (55%
yield). The product obtained was a colorless crystalline
1-Phenyl-2-m-tolyloxy-propan-1-one (3h)
Obtained according to the general procedure from
0.26 g (2.40 mmol) of m-cresol, 0.50 g (2.35 mmol) of
2-bromopropiophenone and 0.58 g (4.23 mmol) of K2CO3
and purified by recrystallization from methanol (54% yield).
The product obtained was a light brown crystalline solid (mp
109-111 °C).1H NMR (600 MHz, CDCl3) d8.07 (d, J7.8 Hz,
2H,Ar-H), 7.57 (t, J 7.8 Hz, 2H,Ar-H), 7.46 (t, J 7.8 Hz, 2H,
Ar-H), 7.10 (t, J 7.9 Hz, 1H, Ar-H), 6.75 (d, J 7.9 Hz, 1H,
Ar-H), 7.10 (t, J 7.9 Hz, 1H, Ar-H), 6.75 (d, J 7.9 Hz, 1H,
Ar-H), 6.72 (s,1H, Ar-H), 6.65 (dd, J1 7.9 Hz, J2 2.4 Hz, 1H,
Ar-H), 5.45 (q, J 7.0 Hz, 1H, CH), 2.27 (s, 3H, CH3), 1.69 (d,
J 7.0 Hz, 3H, CH3); 13C NMR (150 MHz, CDCl3) d 199.2,
157.6, 139.8, 134.4, 133.7, 129.4, 129.0, 128.9, 122.5, 116.3,
112.0, 76.7, 21.6, 18.8. HREIMS m/z 263.1051 [M + Na]+
(calcd. for C16H16NaO2+, 263.1043).
1
solid (mp 145-147 °C). H NMR (250 MHz, DMSO-d6)
d 8.03 (d, J 8.8 Hz, 2H, Ar-H), 8.00 (d, J 8.8 Hz, 2H,
Ar-H), 7.70 (t, J 8.8 Hz, 4H, Ar-H; d, J 15.8 Hz, 1H, CH),
7.50-7.63 (m, 4H,Ar-H), 7.04 (d, J 8.5 Hz, 2H,Ar-H), 6.65
(d, J 15.8 Hz, 1H, CH), 5.68 (s, 2H, CH2), 5.60 (s, 2H, CH2);
13C NMR (62.5 MHz, DMSO-d6) d 194.2, 193.0, 165.9,
160.0, 145.1, 134.3, 134.0, 133.9, 130.2, 128.9, 128.9,
127.9, 127.8, 126.9, 115.1, 114.8, 70.2, 66.4. HREIMS
m/z 401.1389 [M + H]+ (calcd. for C25H21O5+, 401.1384).
(E)-2-Oxo-2-phenylethyl 3-(3-methoxy-4-(2-oxo-2-phenyl-
ethoxy)phenyl)acrylate (4b)
Obtained according to the general procedure from
0.25 g (1.28 mmol) of ferulic acid, 0.50 g (2.51 mmol) of

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Synthesis, Characterization and in vitro Anticancer Activity of Novel 8,4’-Oxyneolignan Analogues
J. Braz. Chem. Soc.
phenacyl bromide and 0.57 g (4.52 mmol) of K2CO3 and
purified by recrystallization from methanol (74% yield).
The product obtained was a white solid (mp 135-137 °C).
1H NMR (600 MHz, CDCl3) d 8.01 (d, J 7.3 Hz, 2H,Ar-H),
7.96 (d, J 7.3 Hz, 2H, Ar-H), 7.73 (d, J 16.1 Hz, 1H, CH),
7.62 (td, J1 7.3 Hz, J2 3.6 Hz, 2H,Ar-H), 7.50 (td, J1 7.8 Hz,
J2 2 Hz, 4H, Ar-H), 7.12 (d, J 1.8 Hz, 1H, Ar-H), 7.06 (dd,
J1 8.4 Hz, J2 1.8 Hz, 1H,Ar-H), 6.80 (d, J 8.4 Hz, 1H,Ar-H),
6.48 (d, J 16.1 Hz, 1H, CH), 5.47 (s, 2H, CH2), 5.40 (s, 2H,
CH2), 3.93 (s, 3H, OCH3); 13C NMR (150 MHz, CDCl3)
d 193.8, 192.4, 166.4, 149.8, 149.7, 145.8, 134.4, 134.4,
134.0, 133.9, 128.9, 128.6, 128.1, 127.8, 122.5, 115.3,
113.9, 110.8, 71.6, 66.0, 56.0. HREIMS m/z 453.1307
[M + Na]+ (calcd. for C26H22NaO6+, 453.1309).
117.7, 117.3, 114.2, 70.9, 66.3. HREIMS m/z 423.1208
[M + Na]+ (calcd. for C25H20NaO5+, 423.1203).
(E)-2-(4-Nitrophenyl)-2-oxoethyl 3-(3-(2-(4-nitrophenyl)-
2-oxoethoxy)phenyl)acrylate (5b)
Obtained according to the general procedure from
0.09 g (0.52 mmol) of 3-hydroxycinnamic acid, 0.25 g
(1.02 mmol) of p-nitrophenacyl bromide and 0.25 g
(1.84 mmol) of K2CO3 and purified by recrystallization
from methanol (45% yield). The product obtained was a
light brown crystalline solid (mp 101-103 °C). 1H NMR
(600 MHz, CDCl3) d 8.20-8.14 (m, 2H, Ar-H), 7.80-7.73
(m, 2H,Ar-H), 7.71-7.62 (m, 2H), 7.56-7.46 (m, 3H,Ar-H),
7.26 (t, J 7.8 Hz, 1H, Ar-H), 7.09 (d, J 7.7 Hz, 1H, Ar-H),
6.85 (s, 1H, Ar-H), 6.82 (d, J 8.4 Hz, 1H, Ar-H), 6.28 (d,
J 16.1 Hz, 1H, CH), 5.12 (s, 2H, CH2), 4.91 (s, 2H, CH2);
13C NMR (150 MHz, CDCl3) d 198.9, 197.4, 165.6, 157.5,
147.3, 146.3, 145.8, 135.5, 134.7, 134.4, 134.4, 134.2,
131.5, 131.4, 130.1, 128.8, 128.8, 124.0, 123.8, 122.2,
117.0, 116.8, 114.1, 72.0, 67.6. HREIMS m/z 513.0901
[M + Na]+ (calcd. for C25H18N2NaO9+, 513.0905).
( E ) - 1 - ( 4 - M e t h ox y p h e ny l ) - 1 - ox o p r o p a n - 2 - y l
3-(4-((1-(4-methoxyphenyl)-1-oxopropan-2-yl)oxy)phenyl)
acrylate (4c)
Obtained according to the general procedure from
0.10 g (0.53 mmol) of p-hydroxycinnamic acid, 0.25 g
(1.03 mmol) of 4-methoxy-8-bromopropiophenone
and 0.26 g (1.85 mmol) of K2CO3 and purified by
recrystallization from methanol (71% yield). The product
obtained was a colorless crystalline solid (mp 142-144 °C).
1H NMR (400 MHz, DMSO-d6) d 8.12 (d, J 8.8 Hz, 2H,
Ar-H), 8.05 (d, J 8.8 Hz, 2H, Ar-H), 7.70 (d, J 8.8 Hz, 2H,
Ar-H), 7.64 (d, J 16.0 Hz, 1H, CH), 7.19-7.08 (m, 4H,
Ar-H), 6.94 (d, J 8.8 Hz, 2H,Ar-H), 6.61 (d, J 16.0 Hz, 1H,
CH), 6.16-6.06 (m, 2H, CH), 3.91 (s, 3H, OCH3), 3.90 (s,
3H, OCH3), 1.59 (d, J 7.0 Hz, 3H, CH3), 1.52 (d, J 7.0 Hz,
3H, CH3); 13C NMR (100 MHz, DMSO-d6) d 196.1, 195.1,
165.8, 163.8, 163.6, 159.2, 144.9, 130.9, 130.8, 130.3,
126.9, 126.7, 126.7, 115.3, 115.0, 114.3, 114.2, 74.3, 71.2,
55.7, 18.5, 17.2. HREIMS m/z 489.1941 [M + H]+ (calcd.
for C29H29O7+, 489.1942).
2-Oxo-2-phenylethyl cinnamate (6a)
Obtained according to the general procedure from
0.38 g (2.56 mmol) of cinnamic acid, 0.50 g (2.51 mmol)
of phenacyl bromide and 0.62 g (4.52 mmol) of K2CO3
and purified by recrystallization from acetone (86% yield).
The product obtained was a colorless crystalline solid
(mp 141-143 °C). 1H NMR (600 MHz, CDCl3) d 7.96 (d,
J 7.2 Hz, 2H, Ar-H), 7.81 (d, J 16.1 Hz, 1H, CH), 7.62 (t,
J 7.2 Hz, 1H, Ar-H), 7.55 (s, 2H, Ar-H), 7.50 (t, J 7.2 Hz,
2H, Ar-H), 7.40 (s, 3H, Ar-H), 6.60 (d, J 16.1 Hz, 1H,
CH), 5.80 (s, 2H, CH2); 13C NMR (150 MHz, CDCl3)
d 192.4, 166.4, 146.3, 134.5, 134.4, 134.0, 130.7, 129.1,
129.0, 128.4, 128.0, 117.2, 66.3. HREIMS m/z 289.0836
[M + Na]+ (calcd. for C17H14NaO3+, 289.0836).
3-[3-(2-Oxo-2-phenyl-ethoxy)-phenyl]-acrylic acid (5a)
Obtained according to the general procedure from
0.42 g (2.56 mmol) 3-hydroxycinnamic acid, 1.00 g
(5.02 mmol) of phenacyl bromide and 1.24 g (9.00 mmol)
of K2CO3 and purified by recrystallization from methanol
(76% yield). The product obtained was a white crystalline
solid (mp 139-141 °C).1H NMR (600 MHz, CDCl3) d 8.02
(d, J 7.3 Hz, 2H, Ar-H), 7.96 (d, J 7.3 Hz, 2H, Ar-H), 7.76
(d, J 16.0 Hz, 1H, CH), 7.59-7.68 (m, 2H,Ar-H), 7.48-7.56
(m, 4H, Ar-H), 7.32 (t, J 8.0 Hz, 1H, Ar-H), 7.19 (d,
J 8.0 Hz, 1H,Ar-H), 7.12 (s, 1H,Ar-H), 7.00 (dd, J1 8.0 Hz,
J2 2.2 Hz, 1H, Ar-H), 6.57 (d, J 16.0 Hz, 1H, CH), 5.47 (s,
2H, CH2), 5.32 (s, 2H, CH2); 13C NMR (150 MHz, CDCl3)
d 194.1, 192.4, 166.3, 158.5, 145.9, 135.9, 134.6, 134.4,
134.1, 134.0, 130.2, 129.0, 129.0, 128.2, 128.0, 122.0,
2-(4-Nitrophenyl)-2-oxoethyl cinnamate (6b)
Obtained according to the general procedure from
0.38 g (2.59 mmol) of cinnamic acid, 0.62 g (2.54 mmol) of
4-nitrophenacyl bromide and 0.63 g (4.56 mmol) of K2CO3
and purified by recrystallization from acetone/hexane (75%
yield). The product obtained was a light yellow crystalline
solid (mp 157-158 °C).1H NMR (600 MHz, CDCl3) d 8.36
(d, J 8.8 Hz, 2H, Ar-H), 8.12 (d, J 8.8 Hz, 2H, Ar-H), 7.81
(d, J 16.0 Hz, 1H, CH), 7.52-7.63 (m, 2H,Ar-H), 7.36-7.45
(m, 3H, Ar-H), 6.58 (d, J 16.0 Hz, 1H, CH), 5.46 (s, 2H,
CH2); 13C NMR (150 MHz, CDCl3) d 191.3, 166.1, 150.7,
146.7, 138.8, 134.1, 130.7, 129.0, 129.0, 128.3, 127.9,
124.1, 116.4, 66.2. HREIMS m/z 334.0696 [M + Na]+
(calcd. for C17H13NNaO5+, 334.0686).

Vol. 28, No. 11, 2017
Souza et al.
2233
1-Oxo-1-phenylpropan-2-yl cinnamate (6c)
1.04 g (4.37 mmol) of 3,4,5-trimethoxycinnamic acid,
1.00 g (4.28 mmol) of 2-bromo-4’-chloroacetophenone
and 1.09 g (7.70 mmol) of K2CO3 and purified by
recrystallization from acetone (45% yield). The product
obtained was a white crystalline solid (mp 117-119 °C).
1H NMR (400 MHz, CDCl3) d 7.90 (d, J 8.0 Hz, 2H,Ar-H),
7.71 (d, J 16.0 Hz, 1H, CH), 7.48-7.46 (m, 2H, Ar-H),
6.78 (s, 2H, Ar-H), 6.50 (d, J 16.0 Hz, 1H, CH), 5.44
(s, 2H, CH2), 3.89 (s, 9H, OCH3); 13C NMR (100 MHz,
CDCl3) d 191.2, 166.1, 153.3, 146.2, 140.3, 140.3, 132.5,
129.6, 129.1, 115.9, 105.4, 65.8, 60.9, 56.1. HREIMS
m/z 391.0948 [M + H]+ (calcd. for C20H20ClO6+, 391.0943).
Obtained according to the general procedure from
0.36 g (2.40 mmol) of cinnamic acid, 0.50 g (2.35 mmol)
of 2-bromopropiophenone and 0.58 g (4.23 mmol) of
K2CO3 and purified by recrystallization from methanol
(57% yield). The product obtained was a white crystalline
solid (mp 70-71 °C). 1H NMR (600 MHz, CDCl3) d 7.99
(d, J 7.2 Hz, 2H, Ar-H), 7.74 (d, J 16.2 Hz, 1H, CH), 7.59
(t, J 7.2 Hz, 1H, Ar-H), 7.51-7.55 (m, 2H, Ar-H), 7.49 (t,
J 7.2 Hz, 2H, Ar-H), 7.37-7.40 (m, 3H, Ar-H), 6.54 (d,
J 16.2 Hz, 1H, CH), 6.12 (q, J 7.0 Hz, 1H, CH), 1.61 (d,
J 7.0 Hz, 3H, CH3); 13C NMR (150 MHz, CDCl3) d 196.9,
166.2, 145.9, 134.5, 134.3, 133.5, 130.5, 128.9, 128.8,
128.5, 128.2, 117.2, 71.4, 17.2. HREIMS m/z 303.0996
[M + Na]+ (calcd. for C18H16NaO3+, 303.0992).
(E)-Methyl-3-(3-methoxy-4-(2-oxo-2-phenylethoxy) phenyl)
acrylate (7a)
Obtained according to the general procedure from
0.75 g (3.59 mmol) of methyl ferulate, 0.70 g (3.52 mmol)
of phenacyl bromide and 0.88 g (6.34 mmol) of K2CO3
and purified by recrystallization from ethanol (53% yield).
The product obtained was a colorless crystalline solid
(mp 105-107 °C). 1H NMR (500 MHz, CDCl3) d 8.00 (d,
J 8.0 Hz, 2H, Ar-H), 7.63-7.59 (m, 2H, Ar-H/CH), 7.50 (t,
J 8.0 Hz, 2H, Ar-H), 7.07 (s, 1H, Ar-H), 7.02 (d, J 8.0 Hz,
1H,Ar-H), 6.77 (d, J 8.0 Hz, 1H,Ar-H), 6.31 (d, J 16.0 Hz,
1H, CH), 5.41 (s, 2H, CH2), 3.92 (s, 3H, OCH3), 3.79 (s,
3H, OCH3); 13C NMR (125 MHz, CDCl3) d 193.7, 167.5,
149.5, 149.3, 144.5, 134.2, 133.9, 128.8, 128.5, 127.9,
122.0, 115.9, 113.6, 110.4, 71.3, 55.9, 51.6. HREIMS
m/z 327.1233 [M + H]+ (calcd. for C19H19O5+, 327.1227).
2-(4-Chlorophenyl)-2-oxoethyl cinnamate (6d)
Obtained according to the general procedure from
0.34 g (2.32 mmol) of cinnamic acid, 0.53 g (2.27 mmol) of
2-bromo-4’-chloroacetophenone and 0.57 g (4.10 mmol) of
K2CO3 and purified by recrystallization from acetone (59%
yield). The product obtained was a white crystalline solid
(mp 128-130 °C). 1H NMR (400 MHz, CDCl3) d 7.89 (d,
J 8.6 Hz, 2H,Ar-H), 7.80 (d, J 16.0 Hz, 1H, CH), 7.52-7.56
(m, 2H, Ar-H), 7.46 (d, J 8.6 Hz, 2H, Ar-H), 7.38-7.41
(m, 3H, Ar-H), 6.58 (d, J 16.0 Hz, 1H, CH), 5.42 (s, 2H,
CH2); 13C NMR (100 MHz, CDCl3) d 191.2, 166.1, 146.2,
140.3, 134.1, 132.5, 130.5, 129.2, 128.9, 128.5, 128.2,
116.7, 65.9. HREIMS m/z 323.0451 [M + Na]+ (calcd. for
C17H13ClNaO3+, 323.0446).
3-{4-[2-(4-Bromo-phenyl)-2-oxo-ethoxy]-3-methoxy-phenyl}-
acrylic acid methyl ester (7b)
(E)-2-Oxo-2-phenylethyl-3-(benzo[d][1,3]dioxol-5-yl)
acrylate (6e)
Obtained according to the general procedure from
0.19 g (0.92 mmol) of methyl ferulate, 0.25 g (0.90 mmol)
of 2,4’-dibromoacetophenone and 0.22 g (1.62 mmol) of
K2CO3 and purified by recrystallization from acetone/
CH2Cl2 (68% yield). The product obtained was a light
yellow crystalline solid (mp 160-162 °C). 1H NMR
(600 MHz, CDCl3) d 7.88 (d, J 8.4 Hz, 2H, Ar-H), 7.64
(d, J 8.4 Hz, 2H, Ar-H), 7.61 (d, J 16.2 Hz, 1H, CH), 7.07
(d, J 1.8 Hz, 1H, Ar-H), 7.03 (dd, J1 8.4 Hz, J2 1.8 Hz, 1H,
Ar-H), 6.79 (d, J 8.4 Hz, 1H,Ar-H), 6.31 (d, J 16.2 Hz, 1H,
CH), 5.31 (s, 2H, CH2), 3.91 (s, 3H, CH3), 3.79 (s, 3H, CH3);
13C NMR (150 MHz, CDCl3) d 193.3, 167.5, 149.8, 149.2,
144.4, 133.1, 132.2, 129.7, 129.2, 128.9, 122.0, 116.3,
114.1, 110.7, 71.8, 56.0, 51.6. HREIMS m/z 427.0158
[M + Na]+ (calcd. for C19H17BrNaO5+, 427.0152).
Obtained according to the general procedure from 0.49 g
(2.56 mmol) of 3,4-(methylenedioxy)cinnamic acid, 0.50 g
(2.51 mmol) of phenacyl bromide and 0.59 g (4.52 mmol) of
K2CO3 and purified by recrystallization from acetone/hexane
(69% yield). The product obtained was a colorless crystalline
solid (mp 144-146 °C). 1H NMR (250 MHz, CDCl3) d
7.95 (d, J 7.4 Hz, 2H, Ar-H), 7.71 (d, J 16.0 Hz, 1H, CH),
7.61 (t, J 7.4 Hz, 1H, Ar-H), 7.49 (t, J 7.4 Hz, 2H, Ar-H),
7.06-7.02 (m, 2H, Ar-H), 6.81 (d, J 7.9 Hz, 1H, Ar-H), 6.42
(d, J 16.0 Hz, 1H, CH), 6.01 (s, 2H, OCH2O), 5.46 (s, 2H,
CH2); 13C NMR (62.5 MHz, CDCl3) d 192.4, 166.4, 149.8,
148.3, 145.8, 134.3, 133.8, 129.2, 128.8, 127.8, 124.7, 114.8,
108.5, 106.6, 101.6, 66.0. HREIMS m/z 349.0478 [M + K]+
(calcd. for C18H14KO5+, 349.0473).
(E)-2-(4-Chlorophenyl)-2-oxoethyl-3-(3,4,5 trimethoxy-
phenyl)acrylate (6f)
(E)-Methyl 3-(3-((1-oxo-1-phenylpropan-2-yl)oxy)phenyl)
acrylate (8)
Obtained according to the general procedure from
Obtained according to the general procedure from

2234
Synthesis, Characterization and in vitro Anticancer Activity of Novel 8,4’-Oxyneolignan Analogues
J. Braz. Chem. Soc.
0.22 g (1.22 mmol) of methyl 3-(3-hydroxyphenyl)acrylate,
0.25 g (1.20 mmol) of 2-bromopropiophenone and 0.30 g
(2.16 mmol) of K2CO3 and purified by recrystallization
from acetone/CH2Cl2 (88% yield). The product obtained
was a light yellow crystalline solid (mp 115-117 °C).
1H NMR (600 MHz, CDCl3) d 8.06 (d, J 7.2 Hz, 2H,Ar-H),
7.63-7.53 (m, 1H, Ar-H/CH), 7.48 (t, J 7.2 Hz, 2H, Ar-H),
7.24 (t, J 7.8 Hz, 1H, Ar-H), 7.10 (d, J 7.8 Hz, 1H, Ar-H),
7.02 (s, 1H,Ar-H), 6.87 (dd, J1 7.8 Hz, J2 1.8 Hz, 1H,Ar-H),
6.34 (d, J 15.6 Hz, 1H, CH), 5.50 (q, J 7.2 Hz, 1H, CH),
3.79 (s, 3H, OCH3), 1.73 (d, J 7.2 Hz, 3H, CH3); 13C NMR
(150 MHz, CDCl3) d 198.5, 167.2, 157.8, 144.4, 135.9,
134.1, 133.8, 130.0, 128.8, 128.8, 121.3, 118.3, 116.9,
114.7, 76.7, 51.7, 18.7. HREIMS m/z 333.1099 [M + Na]+
(calcd. for C19H18NaO4+, 333.1098).
In vitro cell viability assay - MTT assay
All steps in this assay were automated in Liquid
Handling Workstation epMotion® 5070 (Eppendorf,
Vaudaux, Schonenbuch, Switzerland). Cells were
distributed in 96 wells (100 μL cells well-1) and incubated
for 48 h, before addition of test compounds. Cells were
then exposed to the compounds at a concentration of
10 μM. After 24 h of exposure at 37 °C, cell viability was
determined by colorimetric MTT35 (3-(4,5-dimethylthiazol-
2-yl)-2,5-diphenyltetrazoliumbromide) based on the
metabolic ability of active cells to convert the yellow
MTT reagent into a blue insoluble salt (formazan), which
is spectrophotometrically measured. Then, the amount of
formazan produced was dissolved in solution containing
150 μL of isopropanol and optical density was read by a
spectrophotometer at 570 nm (Bio-Tek Power Wave XS).
Absorbance of wells containing the compounds and those
with cells in control (cells treated with vehicle, 0.1%
DMSO) were compared to estimate the cell viability. The
results were expressed as inhibition percentage relative to
control (considered as 100%) and doxorubicin was used
as a reference drug. Compound 6c was also evaluated for
cytotoxicity against human leukemia cells, using the MTT
method. Doxorubicin was used as positive control. All
assays were performed in triplicate and mean standard
deviation (SD) values were used to estimate cell viability.
2-Oxo-2-phenylethyl-4-(2-oxo-2-phenylethoxy)benzoate (9)
Obtained according to the general procedure from
0.35 g (2.56 mmol) p-hydroxybenzoic acid, 0.50 g
(2.51 mmol) of phenacyl bromide and 0.62 g (4.52 mmol)
of K2CO3 and purified by recrystallization from ethanol
(41% yield).The product obtained was a white crystalline
1
solid (mp 146-148 °C). H NMR (500 MHz, CDCl3) d
8.07 (d, J 8.9 Hz, 2H, Ar-H), 7.98 (d, J 8.5 Hz, 2H, Ar-H),
7.94 (d, J 8.5 Hz, 2H, Ar-H), 7.46-7.52 (m, 4H, Ar-H),
6.97 (d, J 8.9 Hz, 2H, Ar-H), 5.54 (s, 2H, CH2), 5.37
(s, 2H, CH2); 13C NMR (100 MHz, DMSO-d6) d 193.6,
192.3, 165.5, 162.0, 134.4, 134.3, 134.1, 133.8, 132.1,
128.9, 128.9, 128.1, 127.8, 122.7, 114.5, 70.5, 66.3.
HREIMS m/z 397.1052 [M + Na]+ (calcd. for C23H18NaO5+,
397.1047).
Results and Discussion
Chemistry
In this work, 25 oxyneolignan analogues (β-ketoethers
or β-ketoesters) were synthesized and the synthetic
strategies for their preparation are summarized in
Scheme 1. The β-ketoethers and β-ketoesters analogues
3a-3j, 4a-4c, 5a-5b, 6a-6f, 7a-7b, 8 and 9 (Table 1) were
obtained following a procedure described by Barata et al.26
Initially, the α-bromoketone intermediates 2a-2b
were prepared according to a known procedure,26 then,
without any purification due to their lacrimogenic property,
were reacted with an in situ generated phenoxy and/or
carboxylate ion. These condensation reactions were carried
out in the presence of K2CO3 and butanone as solvent
furnishing the products in yields ranging from 40 to 100%.
The phenolic/carboxylic compounds were used in an excess
of 2% instead of using excess of α-bromoketone. During
isolation, the volume of solvent was reduced to 1/3 of the
initial volume before work-up, since the slight solubility
of the solvents in water hampers the isolation procedure.
In order to obtain compounds 7a-7b and 8, previous
esterification of the cinnamic acid derivatives with
Biology
Cell culture and treatment
Human myeloid leukemia (KG-1, K-562, HL-60, NB4),
human burkit lymphoma (RAMOS, RAJI), human lymphoid
T leukemia (JURKAT, CEM, MOLT-4), human lymphoid B
leukemia (NALM-6, SUP-B15, RS4;11), human prostatic
adenocarcinoma (PC3, LNCaP), human ovarian carcinoma
(NCI/ADR), human malignant neoplasm cervix uteri
(HeLa), human breast adenocarcinoma (MCF-7), human
osteosarcoma (HOS, U-2 0S, MG-63), human glioblastoma-
astrocytoma, epithelial-like (U-87 MG), human glioblastoma
cell lines (U-138 MG) and human non-small cell lung
cancer (NCI-H1299) were cultured in RPMI medium
supplemented with 10% fetal calf serum (Gibco 16000-044),
1% penicillin (10,000 IU mL-1) and streptomycin 10 mg mL-1
(15,070 Gibco) and exposed to concentration of drugs
(10 μM) in DMSO (0.1%) and maintained at 37 °C in 95%
humidified atmosphere, containing 5% CO2.

Vol. 28, No. 11, 2017
Souza et al.
2235
Scheme 1. Synthesis of 8,4’-oxyneolignan analogues. Reagents and conditions: (i) Br2, CHCl3 r.t. 2 h;(ii) ArOH, K2CO3, butanone or MeCN, 80 °C, 12 h;
(iii) cinnamic acids, K2CO3, butanone or MeCN, 80 °C; (iv) methyl ferulate, butanone or MeCN, 80 °C, 12 h; (v) methyl 3-hydroxycinnamate, K2CO3,
butanone or MeCN, 80 °C; (vi) 4-hydroxy-benzoic acid, K2CO3, butanone, 80 °C.
methanol and sulfuric acid was mandatory to avoid the
competitive nucleophilic displacement of the bromine
atom by the carboxylate anions. This was confirmed in
the obtention of compounds 4a-4c and 5a-5b by using
2 equivalents of 2a-2b. As far as we know, this creates a
new subclass of 8,4’-oxyneolignan analogues, which we
called 8,4’-oxyneolignan cinnamic analogues. Compound 9
was synthesized in a similar fashion.

2236
Synthesis, Characterization and in vitro Anticancer Activity of Novel 8,4’-Oxyneolignan Analogues
J. Braz. Chem. Soc.
Table 1. Chemical structures, yields, and melting points (mp) of the synthesized compounds
Compound
Structure
Yield / %
84
mp / °C
148-150
3a
3b
3c
42
91
74-75
78-80
3d
3e
3f
100
125-127
70-72
-
69
54
3g
3h
3i
66
54
39
111-112
109-111
99-101

Vol. 28, No. 11, 2017
Souza et al.
2237
Table 1. Chemical structures, yields, and melting points (mp) of the synthesized compounds (cont.)
Compound
Structure
Yield / %
69
mp / °C
91-93
3j
4a
4b
4c
55
74
71
145-147
135-137
142-144
5a
5b
76
45
139-141
101-103
6a
6b
6c
86
75
57
141-143
157-158
70-71

2238
Synthesis, Characterization and in vitro Anticancer Activity of Novel 8,4’-Oxyneolignan Analogues
J. Braz. Chem. Soc.
Table 1. Chemical structures, yields, and melting points (mp) of the synthesized compounds (cont.)
Compound
Structure
Yield / %
59
mp / °C
128-130
6d
6e
6f
69
45
144-146
117-119
7a
53
105-107
7b
68
88
41
160-162
115-117
146-148
8
9
1
The H NMR spectra of the products show the
presence of one peak (compounds 3, 6, 7 and 8) or two
peaks (compounds 4, 5 and 9) in between 5.12-6.14 ppm,
assignable to the CH2/CH carbinolic protons. These
signals confirm the formation of the C-O-C bond and,
consequently, the expected products.
Biology
To evaluate the antiproliferative activity, the amount of
surviving cells at the dose level of 10 μM was measured
after 24 h of incubation by the MTT method colorimetric
assay35 and the results were expressed as percentage
of inhibition relative to control and compared with the
reference drug (doxorubicin). The tests were carried out in
triplicate, using doxorubicin as positive control and these
data are schematically listed in Table 2.
All synthesized substances were purified by
crystallization and fully characterized by usual spectroscopic
methods (melting points, HRMS, 1H and 13C NMR). The
chemical structures are described in Table 1.

Vol. 28, No. 11, 2017
Souza et al.
2239
Table 2. Evaluation of cytotoxicity towards leukemia cells (% inhibition)a
for all synthesized compounds
to be evaluated for their activity on cell proliferation on
other nineteen different kinds of human neoplasms, among
tumors and leukemia cell lines. These tests were performed
by using the MTT method, as previously mentioned. The
cells used in this evaluation are listed in the Experimental
section and the obtained results are shown on Tables 3-4
and Figures 1-2.
entry
1
Compound
K-562
11.8
15.6
20.4
28.3
NT
HL-60
15.4
4.8
Nalm-6
< 5
Ramos
14.7
21.0
< 5
3a
2
3b
5.8
3
3c
10.9
26.1
NT
8.7
4
3d
13.4
NT
29.8
NT
5
3e
Table 3. Evaluation of antiproliferative activity towards tumor cells
(% inhibition at 10 μM) for compounds 4a, 4b, 6c, 6d and 9
6
3f
17.2
34.4
13.9
10.3
27.4
34.1
< 10
NT
1.4
< 5
15.0
18.6
10.4
18.5
17.6
48.6
60.4
NT
7
3g
6.5
< 5
Antiproliferative activity / (% inhibition at 10 μM)
Tumor
8
3h
16.9
8.7
< 5
4a
4b
6c
6d
9
Doxorubicin
9
3i
14.1
< 5
PC3
82.9
64.3
76.2
39.7
61.3
72.2
75.7
76.3
21.2
71.7
73.4
67.3
67.6
56.5
35.2
71.1
34.2
5.4
57.1
74.9
60.1
59.0
36.3
43.4
29.8
68.3
41.9
70.2
15.0
47.1
50.6
42.6
31.2
51.4
32.1
27.0
65.0
35.2
36.9
23.1
47.5
50.0
58.4
36.5
33.7
50.0
53.0
27.5
20.9
54.0
35.5
85.1
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
3j
38.1
52.1
57.5
NT
LNCaP
NCI/ADR
MCF-7
HeLa
86.0
4a
56.7
62.0
NT
94.0
4b
72.1
4c
93.7
5a
21.6
12.2
39.3
27.8
52.2
< 5
< 5
12.4
8.6
17.9
15.7
43.7
42.8
84.6
61.3
38.3
36.9
NT
U-87
59.0
5b
< 5
U-138 MG
HOS
73.2
6a
34.1
7.1
32.6
21.7
73.2
56.9
38.5
34.9
NT
66.6
52.0
59.4
24.8
91.5
6b
U-2 OS
MG-63
H1299
90.5
6c
86.3
51.4
33.2
33.7
NT
6d
74.5
6e
43.6
19.9
NT
65.4
6f
7a
Table 4. Evaluation of cytotoxicity towards leukemia cells (% inhibition
at 10 μM) for compounds 4a, 4b, 6c, 6d and 9
7b
13.7
12.8
47.0
71.1
16.3
14.8
66.7
97.7
< 5
9.9
8
9.9
21.7
54.1
81.3
Cytotoxicity / (% inhibition at 10 μM)
Leukemia
cell
9
51.6
65.4
4a
4b
6c
6d
9
Doxorubicin
86.9
doxorubicin
KG-1
55.1
33.2
19.6
56.8
52.4
14.1
48.5
53.0
92.0
48.9
37.9
62.2
61.7
41.5
57.4
40.5
89.7
81.4
65.5
91.2
91.1
87.1
64.4
89.6
34.0
35.2
46.3
51.9
45.6
34.0
51.7
35.8
< 5
aInhibition percentages measured at a single concentration of 10 μM.
NT: not tested.
NB4
45.0
37.1
40.8
57.8
62.0
56.4
50.5
88.2
RAJI
90.8
JURKAT
CEM
97.5
The antiproliferative screening results show that
five compounds presented a promising antiproliferative
activity (≥ 50% of cell inhibition) against the leukemia cell
lines. The other analogues were less active or completely
inactive at the dose of 10 μM. Compounds 4a, 4b, 6d and
9 (entries 11, 12, 19 and 25, respectively) demonstrate
similar inhibition of growth proliferation against HL-60,
Ramos and Nalm-6 (48.6-66.7%). The same substances did
not reveal a good inhibition level against K-562 cells. The
best compound of the series was 6c (entry 18) inhibiting
cell proliferation over 50% against four leukemia cell lines
and very comparable to the positive control used in this test
(entry 26). For Nalm-6 and Ramos cells, the percentage
of inhibition was even superior to doxorubicin inhibition,
presenting 73.2 and 84.6%, respectively.
98.4
MOLT-4
SUP-B15
RS4;11
90.0
89.4
95.2
As can be seen from Table 3 and Figures 1-2, all
compounds exhibited some degree of activity against
the eleven tumor cells used in this assay. β-Ketoester 6c
presented a good antiproliferative activity for tumors,
specially LNCaP (74.9%), HOS (68.3%) and MG-63
(70.2%), whereas compound 6d showed an inhibition rate
greater than 50% only for LNCaP (50.6%), HeLa (51.4%)
and HOS (65.0%) tumors. The oxyneolignan analogue 9
demonstrated a considerable reduction of cell growth on
four tumor cells: LNCaP (50.0%), NCI/ADR (58.4%),
U-138 MG (53.0%) and MG-63 (54.0%).
Compounds which presented significant activity profile
against leukemia cells (4a, 4b, 6c, 6d, 9) were selected

2240
Synthesis, Characterization and in vitro Anticancer Activity of Novel 8,4’-Oxyneolignan Analogues
J. Braz. Chem. Soc.
Figure 1. MTT assay for human tumor cell lines of compounds 6c, 6d and 9.
Figure 2. MTT assay for human tumor cell lines of compounds 4a and 4b.
8,4’-Oxyneolignan cinnamic analogues 4a and 4b were
shown to be active against most cells, exhibiting a good
inhibition profile. The most active compound was 4a,
showing a significant inhibition rate of cell growth against
PC3 (82.9%), MG-63 (71.7%), U-87 (72.2%), U-138 MG
(75.7%) and H1299 (73.4%) tumors. In these last three
cell lines results, the activity of the oxyneolignan 4a was
even superior to that of doxorubicin, the positive control
used in this trial.
The evaluation of compounds 4a, 4b, 6c, 6d and 9 on
leukemic cells are expressed in Table 4 and Figures 3-4.
Compound 9 was most active against CEM, SUP-B15,
MOLT-4 and RS4;11 cells, reducing its growth by 50.5 to
62.0%. Compound 4a presented levels of cell inhibition
against KG-1, JUKART, CEM, RS4;11 around 52.4-56.8%,
whereas a great selectivity and stronger antiproliferative
activity were observed for compound 4b, which exhibited
92.0% of inhibition against KG-1 cells.
Compound 6c was found to be the most promising of
the series, presenting antileukemic activity higher than
50% for all cell lines used in this study and superior or
comparable results when compared to the reference drug.
In the case of KG-1 cells, the extent of inhibition levels
of products 4b and 6c were even better than doxorubicin,
indicating its possible efficacy against this leukemia type.
Considering the results obtained in the antiproliferative
tests (Table 4, Figure 4), in vitro cytotoxicity assay was used
to assess the activity of the most potent compound (6c).
The analogue was tested against eleven human leukemic
cell lines. The assays were carried out in triplicate, and
doxorubicin was used as the positive control. The biological
endpoint was determined according to the concentration,

Vol. 28, No. 11, 2017
Souza et al.
2241
Figure 3. MTT assay for human leukemia cell lines of compounds 4a, 6d and 9.
Figure 4. MTT assay for human leukemia cell lines of compounds 4b and 6c.
which causes fifty percent of cell growth inhibition (IC50).
Compound 6c showed similar values of IC50 against
leukemia cells (Table 5), ranging from moderate to good
activity, except for K-562 cells, for which the drug showed
no activity. Compound 6c induced better cytotoxic effects
on Ramos cells, which presented an IC50 of 9.4 μM.
The correlation between the structures of the synthesized
compounds and their antiproliferative activities leads to the
conclusion that the compounds bearing a cinnamic moiety
on its structure possess better potency in MTT assay. The
presence of the cinnamic portion in the basic structure of
the 8,4’-oxyneolignans and the variation on its position on
the aromatic ring affect considerably the antiproliferative
activities. When comparing compounds 4a-4b and 5a-5b,
7a-7b and 8 better inhibition showed by compounds 4a
and 4b against cancer cells could be attributed to its para
substitution, whereas meta-substituted compounds 5a-5b
did not show an expressive percentage of inhibition.
However, in compounds 7a-7b and 8, the cinnamate moiety
and the absence of an aromatic ring may be responsible for
the drastic decrease in activity.
In the 8,4’-oxyneolignans 3a-3j, the effects of
substitution on the aromatic rings were not clearly
observed. These compounds did not show enhanced activity
when comparing substituent groups, positions and side
chain length.
Among compounds 6a-6f, as shown in Tables 2 and 3,
the effect of alkyl chain substitution in compound 6c greatly

2242
Synthesis, Characterization and in vitro Anticancer Activity of Novel 8,4’-Oxyneolignan Analogues
J. Braz. Chem. Soc.
Table 5. In vitro cytotoxicity of compound 6c against leukemia cell lines
Acknowledgments
Leukemia cell
K-562
Doxorubicin IC50 / μM
6c IC50 / μM
> 100
12.0
The authors gratefully acknowledge National Council
for Scientific and Technological Development (CNPq) and
University of Brasilia (UnB) for financial support.
3.1
0.8
0.5
0.4
5.7
1.0
0.2
0.5
0.1
0.5
0.1
NT
KG-1
HL-60
12.6
NB4
12.6
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Ramos
RAJI
9.4
18.5
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Submitted: February 17, 2017
Published online: April 26, 2017