Vasoactive thiomethyl-pyrimidines: Promising drug candidates with vascular activity

Article abstract of DOI:10.21577/0103-5053.20160289

Pyrimidines and their derivatives are present in various biologically active molecules. Most of the synthetic methods employed to achieve the pyrimidinone ring consist of two stages: The synthesis of a Michael intermediate from an aldehyde and an "active

Full text of DOI:10.21577/0103-5053.20160289

http://dx.doi.org/10.21577/0103-5053.20160289
J. Braz. Chem. Soc., Vol. 28, No. 7, 1266-1273, 2017.
Printed in Brazil - ©2017 Sociedade Brasileira de Química
0103 - 5053 $6.00+0.00
Article
Vasoactive Thiomethyl-Pyrimidines: Promising Drug Candidates with
Vascular Activity
Audrey N. de Andrade,a Alice V. Araújo,b Hugo B. W. Barbosa,a Almir G. Wanderley,c
Oscar L. Malta*,a and Janaína V. dos Anjos*,a
aDepartamento de Química Fundamental, Universidade Federal de Pernambuco,
50740-560 Recife-PE, Brazil
bNúcleo de Nutrição, Universidade Federal de Pernambuco,
55608-680 Vitória de Santo Antão-PE, Brazil
cDepartamento de Fisiologia e Farmacologia, Universidade Federal de Pernambuco,
50670-901 Recife-PE, Brazil
Pyrimidines and their derivatives are present in various biologically active molecules. Most of
the synthetic methods employed to achieve the pyrimidinone ring consist of two stages: the synthesis
of a Michael intermediate from an aldehyde and an “active methylene” containing compound; and
the condensation of this intermediate with a molecule containing an uranium moiety. This may take
one to two days of laboratory work. In this paper we describe a new methodology in which these
derivatives are obtained via multicomponent synthesis mediated by ultrasound in only 2 hours.
In order to obtain water-soluble pyrimidinone derivatives, our previous compounds were further
converted into their sodium salts. In pharmacologic studies, these salts inhibited phenylephrine-
induced contraction in isolated rat aorta, suggesting that they may act as alpha-1 antagonists and,
therefore, are candidates for anti-hypertensive drugs.
Keywords: pyrimidines, vascular activity, hypertension, drug discovery, ultrasound
Introduction
When an alpha-1 adrenergic receptor is blocked by an
antagonist, it inhibits the contraction of blood vessels and
vasodilation takes place. This results in blood pressure drop.
For this reason, alpha-1 adrenergic receptor antagonists
are mainly used as anti-hypertensive drugs.5 These drugs
include Prazosin (I), Terazosin (II) and Doxazosin (III)
(Figure 1), that are quite similar in their structures,
belonging to the 2,4-diamino-6,7-dimethoxyquinazolines
class of AR-1 receptor antagonists.1,4
Alpha adrenergic receptors are involved in many
biological actions of catecholamines. Therefore, those
receptors have been explored as targets for a large
number of substances and they have been employed for
the treatment of several pathologies. The most important
effects of alpha adrenergic antagonists are related to the
cardiovascular system.1,2
The catecholamines receptors can be classified
according to their binding specificities, formerly alpha-1
and alpha-2.1 They are quite similar and belong to G
protein-coupled superfamily receptors.1,3 While alpha-1
adrenergic receptors mediate catecholamine-induced
peripheral vasoconstriction, alpha-2 is mainly involved in
the peripheral and central regulation of the sympathetic
nervous system.Alpha-1 receptors can also be divided into
three subtypes, alpha 1A, alpha 1B, and alpha 1D, that have
distinct actions and tissue expression.3,4
ItcanbeobservedthatPrazosin,TerazosinandDoxazosin
have a pyrimidine ring in their chemical structures.4 In
fact, pyrimidines and their oxo-derivatives are described
in the literature as presenting a wide array of biological
activities such as antitumor,6,7 interferon inducer,8 antiviral,9
anti-hipertensive,10 hypoglycemic,11 anticonvulsant,12
antinociceptive,13,14 and anti-inflammatory.15
Pyrimidine or pyrimidinone rings can be synthesized in
several ways.16 Most often, for the synthesis of these type of
heterocycles, a Michael adduct and a uronium-containing
molecule (guanidine,17 amidines,18 urea,19 thiourea and
their derivatives20) react in the presence of an organic base.
*e-mail: omlmalta@gmail.com; janaina.anjos@ufpe.br

Vol. 28, No. 7, 2017
de Andrade et al.
1267
Figure 1. Molecular structures of Prazosin (I), Terazosin (II) and Doxazosin (III).
This reaction results in an alicyclic compound that can
be converted into the heterocycles of interest employing
cyclization conditions. This whole process may take
two days of laboratory work for the preparation of one
product.9-11,14,18 These findings encouraged us to search
new ways to perform these reactions and to evaluate the
biological activity of these heterocyclic derivatives.
Multicomponent reactions (MCRs) are procedures in
which three or more reagents are mixed together to yield
a product, in a domino process.21 These types of reaction
can be efficiently accomplished by employing ultrasonic
irradiation.22,23 The use of ultrasound energy in organic
reactions relies on the cavitation effect, which induces
mass transfer enhancement, leading to better reaction
yields and shorter reaction times. Using mild conditions
and sonochemistry, a lot of heterocyclic rings have been
synthesized, including pyrimidines and their derivatives.24-27
As an example, Pagadala et al.28 have used ultrasound
irradiation to synthesize pyrimidines in aqueous NaOH
solution, by addition of aromatic aldehyde to acetyl acetone
and thiourea, via multicomponent approach.
containing GF254 (E. Merck). The reactions were carried
out in a Branson B2510E-DTH ultrasonic bath operating
at frequency of 40 kHz. Melting points were determined on
a Büchi apparatus and are uncorrected. Nuclear magnetic
resonance (NMR) spectra were recorded with a Varian
UNMRS 400 MHz spectrometer. Infrared spectra were
obtained on a Brucker IFS66 FT-IR, using KBr pellets.
Exact mass measurements of the molecular ions were
obtained on a Shimadzu Electrospray LC/MS-IT-TOF.
An elemental analysis was performed with a Carlo Erba
instrument model E-1110.
Multicomponent synthesis of 2-methylsulfanyl-6-oxo-4-aryl-
1,6-dihydro-pyrimidine 5-carbonitriles (4a-j)
To an Erlenmeyer (capacity: 50 mL), were added
1.2 mmol (108 mg) of 2-methyl-2-thiopseudourea
hemisulfate and 2 mmol (276 mg) of potassium carbonate
dissolved in 4 mL of distilled water. This mixture was
sonicated at room temperature until the neutralization of
the 2-methyl-2-thiopseudourea salt. To a beaker (capacity:
20 mL), 1 mmol of the corresponding aromatic aldehyde
and 1.2 mmol (136 mg, 0.13 mL) of ethyl cyanoacetate were
dissolved in 4 mL of ethanol. This solution was added to
the Erlenmeyer containing the 2-methyl-2-thiopseudourea
basic solution. The reaction mixture was sonicated at a
frequency of 40 kHz at 50 °C for 2 hours. The product was
precipitated by addition of concentrated HCl and poured
into ice. The formed precipitate was then vacuum filtered,
washed with distilled water (50 mL), and recrystallized
from ethanol to provide the pure pyrimidinones.
These positive impacts on MCRs by the use of
ultrasound irradiation have prompted us to investigate
and to compare the methodologies (sonochemistry and
conventional stirring) in the synthesis of thiomethyl-
pyrimidinones. This study also aimed to evaluate whether
the synthesized compounds have alpha-1 antagonist action
in vascular reactivity studies.
Experimental
General information
2-Methylsulfanyl-6-oxo-4-phenyl-1,6-dihydro-pyrimidine-
5-carbonitrile (4a)
White crystals; yield 72%; m.p. 280-282 °C (EtOH);
Rf 0.84 (9:1 CHCl3/MeOH v/v); IR (KBr pellet) νmax / cm-1
All commercially available reagents were used without
any further purification and the reactions were monitored by
thin layer chromatography (TLC) analysis with TLC plates

1268
Vasoactive Thiomethyl-Pyrimidines
J. Braz. Chem. Soc.
2854 (CHarom), 2220 (C≡N), 1662 (C=O), 1540 (C=N);
1H NMR (400 MHz, DMSO-d6) d 2.61 (s, 3H, SCH3), 7.54-
7.62 (m, 3H, Ph-H), 7.95 (dd, 1H, J 2.4, 8.1 Hz, Ph-H),
7.97 (dd, 1H, J 2.4, 8.1 Hz, Ph-H); 13C NMR (100 MHz,
DMSO-d6) d 13.8, 93.4, 116.4, 129.1, 126.2, 132.2,
135.8, 161.4, 167.1, 167.7; HRMS-ESI (high resolution
mass spectrometry-electrospray ionization) m/z calcd. for
C12H8N3OS [M - H]+: 242.0388, found: 242.0081.
131.6, 134.4, 160.8, 166.0, 166.8; HRMS-ESI m/z calcd.
for C12H8BrKN3OS [M + K]+: 361.2787, found: 361.8388.
2-Methylsulfanyl-6-oxo-4-p-fluorophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile (4f)
White crystals; yield 56%; m.p. 283-285 °C (EtOH);
Rf 0.58 (9:1 CHCl3/MeOH v/v); IR (KBr) νmax / cm-1
2835, 2223, 1655, 1536; 1H NMR (400 MHz, DMSO-d6)
d 2.59 (s, 3H, SCH3), 7.39 (d, 2H, J 8.4 Hz, Ph-H), 8.10
(d, 2H, J 8.4 Hz, Ph-H), 13.76 (s, 1H, NH); 13C NMR
(100 MHz, DMSO-d6) d 13.3, 92.8, 115.8, 115.9, 131.4,
131.5, 131.7, 160.8, 165.3, 165.3, 165.7, 166.6; HRMS-
ESI m/z calcd. for C12H8FN3NaOS [M + Na]+: 284.0270,
found: 283.9578.
2-Methylsulfanyl-6-oxo-4-o-tolyl-1,6-dihydro-pyrimidine-
5-carbonitrile (4b)
White crystals; yield 55%; m.p. 205-207 °C (EtOH);
Rf 0.68 (9:1 CHCl3/MeOH v/v); IR (KBr) νmax / cm-1 2843,
2224, 1665, 1546; 1H NMR (400 MHz, DMSO-d6) d 2.31
(s, 3H, PhCH3), 2.52 (s, 3H, SCH3), 7.30-7.45 (m, 4H,
Ph-H); 13C NMR (100 MHz, DMSO-d6) d 13.0, 19.1, 95.8,
114.9, 125.6, 128.4, 130.0, 130.5, 135.4, 166.6; HRMS-ESI
m/z calcd. for C13H10N3O2S [M - H]+: 256.0545, found:
256.0243.
4-(4-Methoxy-phenyl)-2-methylsulfanyl-6-oxo-1,6-dihydro-
pyrimidine-5-carbonitrile (4g)
White crystals; yield 55%; m.p. 280-283 °C (EtOH);
Rf 0.78 (9:1 CHCl3/MeOH v/v); IR (KBr) νmax / cm-1 2841,
2216, 1650, 1606; 1H NMR (400 MHz, DMSO-d6) d 2.60
(s, 3H, SCH3), 3.85 (s, 3H, PhOCH3), 7.10 (dd, 2H, J 2.5,
9.3 Hz, Ph-H), 7.11 (dd, 2H, J 2.5, 9.3 Hz, Ph-H), 8.02
(dd, 2H, J 2.5, 9.3 Hz, Ph-H), 8.02 (dd, 2H, J 2.5, 9.3 Hz,
Ph-H); 13C NMR (100 MHz, DMSO-d6) d 13.7, 56.0, 91.7,
114.5, 116.9, 127.7, 131.3, 162.7, 166.4, 166.6; HRMS-ESI
m/z calcd. for C13H12N3O2S [M + H]+: 274.0650, found:
274.0305.
2-Methylsulfanyl-6-oxo-4-p-tolyl-1,6-dihydro-pyrimidine-
5-carbonitrile (4c)
White crystals; yield 50%; m.p. 285-288 °C (EtOH);
Rf 0.79 (9:1 CHCl3/MeOH v/v); IR (KBr) νmax / cm-1 2754,
2221, 1652, 1523; 1H NMR (400 MHz, DMSO-d6) d 2.39
(s, 3H, PhCH3), 2.60 (s, 3H, SCH3), 7.37 (d, 2H, J 10.8 Hz,
Ph-H), 7.89 (d, 2H, J 10.8 Hz, Ph-H); 13C NMR (100 MHz,
DMSO-d6) d 13.7, 21.6, 92.8, 116.6, 129.2, 129.6, 132.9,
142.6, 161.4, 166.8, 167.4; HRMS-ESI m/z calcd. for
C13H10N3O2S [M - H]+: 256.0545, found: 256.0245.
2-Methylsulfanyl-6-oxo-4-m-nitrophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile (4h)
White crystals; yield 53%; m.p. 258-263 °C (EtOH);
Rf 0.41 (9:1 CHCl3/MeOH v/v); IR (KBr) νmax / cm-1 2860,
2225, 1669, 1543; 1H NMR (400 MHz, DMSO-d6) d 2.63
(s, 3H, SCH3), 7.88 (dd, 1H, J 7.6 Hz, Ph-H), 8.41 (dd,
2H, J 7.6, 7.6 Hz, Ph-H), 8.74 (s, 1H, Ph-H); 13C NMR
(100 MHz, DMSO-d6) d 13.2, 93.8, 115.3, 123.1, 125.9,
130.3, 134.8, 136.7, 147.7, 160.8, 164.8, 167.5; HRMS-ESI
m/z calcd. for C12H8KN4O3S [M + K]+: 326.9954, found:
326.9211.
2-Methylsulfanyl-6-oxo-4-p-chlorophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile (4d)
White crystals; yield 47%; m.p. 293-295 °C (EtOH);
Rf 0.54 (9:1 CHCl3/MeOH v/v); IR (KBr) νmax / cm-1
2865, 2222, 1671, 1546; 1H NMR (400 MHz, DMSO-d6)
d 2.60 (s, 3H, SCH3), 3.43 (s, 1H, NH), 7.64 (dd, 2H,
J 8.4 Hz, Ph-H), 7.98 (dd, 2H, J 8.4 Hz, Ph-H); 13C NMR
(100 MHz, DMSO-d6) d 13.3, 93.0, 115.8, 128.7,
130.6, 134.1, 136.6, 161.0, 165.9, 167.0; HRMS-ESI
m/z calcd. for C12H8ClN3NaOS [M + Na]+: 299.9974,
found: 299.9821.
2-Methylsulfanyl-6-oxo-4-p-nitrophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile (4i)
Yellow crystals; yield 53%; m.p. 290-293 °C (EtOH);
Rf 0.33 (9:1 CHCl3/MeOH v/v); IR (KBr) νmax / cm-1 3101,
2230, 1696, 1525; 1H NMR (400 MHz, DMSO-d6) d 2.60
(s, 3H, SCH3), 3.35 (s, 3H, PhOCH3), 8.20 (dd, 2H, J 2.5,
8.8 Hz, Ph-H), 8.40 (dd, 2H, J 2.5, 8.8 Hz, Ph-H); 13C NMR
(100 MHz, DMSO-d6) d 13.4, 115.5, 123.2, 129.4, 130.2,
141.3, 149.0, 165.4, 167.6. Anal. calcd. for C12H8N4O3S:
C, 50.00; H, 2.80; N, 19.43; S, 11.12%; found: C, 49.64;
H, 3.08; N, 19.45; S, 10.71%.
2-Methylsulfanyl-6-oxo-4-p-bromophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile (4e)
White crystals; yield 40%; m.p. > 300 °C (EtOH);
Rf 0.75 (9:1 CHCl3/MeOH v/v); IR (KBr) νmax / cm-1
2927, 2215, 1650, 1572; 1H NMR (400 MHz, DMSO-d6)
d 2.61 (s, 3H, SCH3), 7.79 (d, 2H, J 8.4 Hz, Ph-H), 7.91
(d, 2H, J 8.4 Hz, Ph-H), 13.77 (s, 1H, NH); 13C NMR
(100 MHz, DMSO-d6) d 13.3, 93.0, 115.7, 125.5, 130.6,

Vol. 28, No. 7, 2017
de Andrade et al.
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4-(3,4-Dichloro-phenyl)-2-methylsulfanyl-6-oxo-1,6-dihydro-
pyrimidine-5-carbonitrile (4j)
1H NMR (400 MHz, DMSO-d6) d 2.35 (s, 3H, SCH3),
7.52 (d, 2H, J 8.8 Hz, Ph-H), 7.79 (d, 2H, J 8.8 Hz, Ph-H);
13C NMR (100 MHz, DMSO-d6) d 13.3, 88.4, 119.9,
128.17, 129.9, 134.4, 136.5, 165.6, 170.5, 172.64.
White crystals; yield 40%; m.p. 297-300 °C (EtOH);
Rf 0.53 (9:1 CHCl3/MeOH v/v); IR (KBr) νmax / cm-1 2856,
2224, 1666, 1549; 1H NMR (400 MHz, DMSO-d6) d 2.61
(s, 3H, SCH3), 3.35 (s, 1H, NH), 7.84 (d, 1H, J 8.0 Hz,
Ph-H), 7.93 (d, 1H, J 8.0 Hz, Ph-H), 8.15 (s, 1H, Harom);
13C NMR (400 MHz, DMSO-d6) d 13.2, 93.5, 115.3, 128.6,
130.3, 130.8, 131.4, 134.4, 135.6, 160.6, 164.4, 167.2.Anal.
calcd. for C12H7Cl2N3OS: C, 46.17; H, 2.26; N, 13.46; S,
10.27%; found: C, 46.02; H, 2.54; N, 13.38; S, 9.45%.
2-Methylsulfanyl-6-oxo-4-p-bromophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile sodium salt (5e)
White crystals; IR (KBr) νmax / cm-1 3434, 2208, 1546,
1
1481; H NMR (300 MHz, D2O) d 2.51 (s, 3H, SCH3),
7.65 (d, 2H, J 9.0 Hz, Ph-H), 7.71 (d, 2H, J 9.0 Hz, Ph-H);
13C NMR (100 MHz, DMSO-d6) d 13.3, 88.4, 119.9, 123.2,
130.2, 131.1, 136.8, 165.7, 170.58, 172.7.
Synthesis of 2-methylsulfanyl-6-oxo-4-aryl-1,6-dihydro-
pyrimidine 5-carbonitriles sodium salts (5a-j)
2-Methylsulfanyl-6-oxo-4-p-fluorophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile sodium salt (5f)
To an Erlenmeyer (capacity: 50 mL) containing 1 mmol
of the corresponding pyrimidinone derivative (4a-j), were
added 1 mmol of NaOH dissolved in 20 mL of ethanol. The
reaction mixture was sonicated at a frequency of 40 kHz
for 30 minutes. The contents were then evaporated and the
formedprecipitatewasrecrystallizedfromamethanol/diethyl
ether mixture to afford the corresponding sodium salts.
White crystals; IR (KBr) νmax / cm-1 3068, 2204, 1550,
1
1483 (C=N); H NMR (400 MHz, D2O) d 2.52 (s, 3H,
SCH3), 7.24-7.31 (m, 2H, Ph-H), 7.77-7.82 (m, 2H, Ph-H);
13C NMR (100 MHz, DMSO-d6) d 13.3, 88.3, 114.9, 115.1,
120.2, 130.4, 130.5, 134.2, 134.16, 161.6, 164.1, 165.7,
170.5, 172.5.
4-(4-Methoxy-phenyl)-2-methylsulfanyl-6-oxo-1,6-dihydro-
pyrimidine-5-carbonitrile sodium salt (5g)
2-Methylsulfanyl-6-oxo-4-phenyl-1,6-dihydro-pyrimidine-
5-carbonitrile sodium salt (5a)
White crystals; IR (KBr pellet) νmax / cm-1 2217, 1649,
1606; 1H NMR (400 MHz, DMSO-d6) d 2.60 (s, 3H, SCH3),
3.85 (s, 3H, PhOCH3), 7.11 (dd, 2H, J 8.8 Hz, Ph-H), 8.0
(dd, 2H, J 8.8 Hz, Ph-H); 13C NMR (100 MHz, DMSO-d6)
d 13.4, 88.6, 119.8, 128.3, 129.9, 130.5, 130.9, 132.4,
138.2, 164.1, 170.1, 172.7.
White crystals; IR (KBr) νmax / cm-1 2204, 1541, 1490;
1H NMR (400 MHz, D2O) d 2.51 (s, 3H, SCH3), 7.51-7.59
(m, 3H, Ph-H), 7.73 (dd, 2H, J 2.8, 8.4 Hz, Ph-H); 13C NMR
(100 MHz, DMSO-d6) d 13.3, 88.6, 120.1, 128.1, 128.1,
129.7, 137.7, 166.9, 170.6, 172.4.
2-Methylsulfanyl-6-oxo-4-o-tolyl-1,6-dihydro-pyrimidine-
5-carbonitrile sodium salt (5b)
2-Methylsulfanyl-6-oxo-4-m-nitrophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile sodium salt (5h)
White crystals; IR (KBr pellet) νmax / cm-1 2221, 1649,
1528; 1H NMR (400 MHz, DMSO-d6) d2.30 (s, 3H, PhCH3),
2.51 (s, 3H, SCH3), 7.30-7.46 (m, 4H, Ph-H); 13C NMR
(100 MHz, DMSO-d6) d 13.2, 19.3, 95.9, 115.1, 125.7,
128.6, 130.2, 130.7, 135.5, 135.7, 160.4, 166.6, 194.4.
White crystals; IR (KBr) νmax / cm-1 2202, 1528, 1548;
1H NMR (400 MHz, DMSO-d6) d 2.20 (s, 3H, SCH3), 7.59
(dd, 2H, J 8.0, 8.0 Hz, Ph-H), 8.10 (d, 2H, J 8.4 Hz, Ph-H),
8.15 (d, 2H, J 8.4 Hz, Ph-H), 8.44 (s, 1H, Ph-H); 13C NMR
(100 MHz, DMSO-d6) d 13.3, 115.6, 123.3, 126.0 130.4,
134.9, 136.8, 147.7, 164.9, 167.6.
2-Methylsulfanyl-6-oxo-4-p-tolyl-1,6-dihydro-pyrimidine-
5-carbonitrile sodium salt (5c)
2-Methylsulfanyl-6-oxo-4-p-nitrophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile sodium salt (5i)
White crystals; IR (KBr) νmax / cm-1 2923, 2210,
1
1650, 1543; H NMR (400 MHz, DMSO-d6) d 2.38 (s,
Yellow crystals; IR (KBr) νmax / cm-1 2223, 1675, 1527;
1H NMR (300 MHz, D2O) d 2.51 (s, 3H, SCH3), 7.95 (d,
2H, J 8.4 Hz, Ph-H), 8.37 (d, 2H, J 8.4 Hz, Ph-H); 13C NMR
(100 MHz, DMSO-d6) d 13.3, 89.4, 119.3, 123.4, 129.6,
143.7, 148.0, 165.0, 169.7, 172.7.
3H, PhCH3), 2.06 (s, 3H, SCH3), 7.35 (d, 2H, J 8.0 Hz,
Ph-H), 7.88 (d, 2H, J 8.0 Hz, Ph-H); 13C NMR (100 MHz,
DMSO-d6) d 13.2, 21.0, 92.3, 116.0, 128.7, 128.8, 129.1,
132.4, 142.1, 160.9, 166.3, 166.8.
2-Methylsulfanyl-6-oxo-4-p-chlorophenyl-1,6-dihydro-
pyrimidine-5-carbonitrile sodium salt (5d)
4-(3,4-Dichloro-phenyl)-2-methylsulfanyl-6-oxo-1,6-dihydro-
pyrimidine-5-carbonitrile sodium salt (5j)
White crystals; IR (KBr) νmax / cm-1 2200, 1663, 1547;
White crystals; IR (KBr) νmax / cm-1 2202, 1528, 1488;

1270
Vasoactive Thiomethyl-Pyrimidines
J. Braz. Chem. Soc.
1H NMR (400 MHz, DMSO-d6) d 2.61 (s, 3H, SCH3), 7.92
(dd, 1H, J 2.0, 8.0 Hz, Ph-H), 7.94 (dd, 1H, J 2.0, 8.0 Hz,
Ph-H), 8.15 (d, 1H, J 8.0 Hz, Ph-H); 13C NMR (100 MHz,
DMSO-d6) d 13.3, 93.65, 115.6, 128.8, 130.4, 130.5, 130.9,
131.5, 134.4, 135.8, 160.7, 164.7, 167.3.
multiple comparison test. The analysis was performed
using GraphPad Prism 6.0®. The level of significance for
rejection of the null hypothesis was set at 5% (p < 0.05).
Results and Discussion
Vascular reactivity studies
Prior to the synthesis of the 2-methylsulfanyl-6-oxo-
4-aryl-1,6-dihydro-pyrimidine 5-carbonitriles, it was
necessary to choose and to optimize the reaction conditions.
In our synthetic approach, aromatic aldehydes (1a-j) react
with ethyl cyanoacetate (2) and 2-methyl-2-thiopseudourea
hemisulfate (3), in the presence of potassium carbonate as
base. To this end, some experiments were performed, in
which we tested three parameters we believe that could
generate better reaction yields or smaller reaction times:
solvent, method (ultrasound or conventional stirring)
and temperature. These results are depicted in Table 1.
The reaction conditions in those preliminary tests were
optimized with benzaldehyde.
Male Wistar rats (250-300 g) were obtained from the
Department of Physiology and Pharmacology, Federal
University of Pernambuco (UFPE). They were kept under
standard environmental conditions (12 h dark/light cycle)
and temperature (22 2 °C). Water and industrialized dry
food (Labina®, Purina, Brazil) were available ad libitum.All
the experimental protocols were submitted to and approved
by the UFPE’sAnimal Experimentation Ethics Committee,
under license No. 021446/2013-12 in accordance with the
National Institute of Health Guide for the Care and Use of
Laboratory Animals.
The rats were killed by CO2 inhalation. The thoracic
aorta was quickly removed, dissected free, and cut into
4 mm-long rings. The aortic rings were placed between
two stainless-steel stirrups and connected to an isometric
force transducer, in order to measure the tension. The rings
were placed in a 10 mL organ chamber containing Krebs
solution with the following composition (mmol L-1): NaCl
130, KCl 4.7, KH2PO4 1.2, MgSO4 1.2, NaHCO3 14.9,
glucose 5.5, and CaCl2 1.6. The solution was maintained
at pH 7.4, and gassed with 95% O2 and 5% CO2, at 37 °C.
The rings were initially stretched to a basal tension of 1.5 g
and allowed to equilibrate for 60 min in the bath, being
washed every 15 min. The endothelium was mechanically
removed and the effectiveness of the removal was assessed
by the absence of relaxation induced by acetylcholine
(1 μmol L-1) in the presence of contractile tone induced by
phenylephrine (0.1 μmol L-1).
Cumulative concentration-effect curves for
phenylephrine (0.1 nmol L-1-10 μmol L-1) were constructed
in the absence or in the presence of the derivatives or
Prazosin (102 μmol L-1). When the derivatives were
used, they were added to the bath 30 min prior to the
concentration-effect curve for phenylephrine to allow the
equilibrium time necessary for the molecules to bind to
the target receptors.
These results clearly indicate that the best conditions
employed an equal mixture of ethanol and water as
solvents, using ultrasound irradiation at 50 °C, showing
that 4a can be obtained via MCR in 72%, after isolation
and purification. This way, we decided to perform the
synthesis of a series of 2-methylsulfanyl-6-oxo-4-aryl-
1,6-dihydro-pyrimidine 5-carbonitriles, under the same
reaction conditions (Table 2).
There are, possibly, two subsequent reactions occurring
in the reaction media in order to build the heterocycle
scaffold: the first one is the Knoevenagel reaction,29
where an aldehyde reacts with ethyl cyanoacetate, to form
Michael’s intermediate. This latter can now react with
2-methyl-2-thiopseudourea. Michael’s adduct can then be
attacked by the single electron pair of a nitrogen atom in
the uronium portion. There is then a sequence of additions
with a ring closure to form the pyrimidinone cycle. The last
step in this mechanistic explanation relies on the oxidation
promoted by atmospheric oxygen, resulting on the loss of
a H2 molecule and leading to the heterocycle (Scheme 1).
Once those compounds are closely related to
Prazosin (I),4 we have decided to evaluate their vascular
reactivity in isolated aorta of rats. However, those derivatives
showed poor solubility in water, which could derail the
tests. Therefore, we decided to synthesize the sodium
salts of pyrimidinones 4a-j. The synthesis was performed
by mixing the pyrimidinones in equimolar quantity of
NaOH dissolved in ethanol, under ultrasound irradiation
(Scheme 2). Those derivatives were much more soluble
in water, so we could carry out the pharmacological tests.
Phenylephrine is a selective alpha-1 adrenergic
agonist. When phenylephrine binds to alpha-1 adrenergic
Statistical analysis
The results of the vascular reactivity studies were
expressed as mean standard error of mean (S.E.M.).
Differences between means were determined using one
way analysis of variance (ANOVA) followed by Dunnet’s

Vol. 28, No. 7, 2017
de Andrade et al.
1271
Table 1. Influence of reaction conditions on the synthesis of 2-methylsulfanyl-6-oxo-4-aryl-1,6-dihydro-pyrimidine 5-carbonitrile
entry
Temperature / °C
Method
Solvent
acetonitrile
water
Yielda / %
1
50
50
50
50
50
50
50
r.t
conventional
conventional
conventional
ultrasound
ultrasound
ultrasound
ultrasound
conventional
conventional
conventional
ultrasound
ultrasound
ultrasound
conventional
39
14
29
44
8
2
3
ethanol/water
acetonitrile
water
4
5
6
ethanol
52
72
30
7
7
ethanol/water
acetonitrile
water
8
9
r.t
10
r.t
ethanol/water
acetonitrile
water
36
47
6
11
r.t
12
r.t
13
r.t
ethanol/water
ethanol
40
0
14
r.t
aIsolated yieds.
Table 2. Ultrasound-assisted multicomponent reaction for the synthesis of 6-oxo-2,4-diaryl-1,6-dihydro-pyrimidine-5-carbonitriles (4a-j)
Compound
Ar
Yield / %
72
4a
4b
4c
4d
4e
4f
Ph
o-tolyl
55
p-tolyl
50
p-ClPh
p-BrPh
p-FPh
47
40
56
4g
4h
4i
p-OCH3Ph
m-NO2Ph
p-NO2Ph
3,4-diClPh
55
53
53
4j
40
receptor, it induces a cellular cascade that leads to vascular
smooth muscle cells contraction.1 It can be assessed by
cumulative concentration-effect curves. Rossum and Van
Den Brink30 described the construction of concentration-
effect curves. It is based on the cumulative addition of
crescent concentrations of a drug to a tissue in an organ
bath. From the concentration versus effect graphic, the
pharmacological parameter “maximum effect” may be
calculated. This parameter represents the maximum effect
a drug can generate over a specific tissue.
Pharmacologic antagonists are substances that bind to
the receptor and do not induce receptor activation (effect),

1272
Vasoactive Thiomethyl-Pyrimidines
J. Braz. Chem. Soc.
Scheme 1. Reaction mechanism for the formation of thiomethyl-pyrimidinones.
Scheme 2. Thiomethyl-pyrimidinone salts preparation.
but prevent the agonist from binding with the receptor and
inducing its cellular effect. Therefore, in the presence of an
antagonist, the effect of the agonist is reduced or, depending
on the concentration, even absent. Prazosin, Terazosin
and Doxazosin are examples of alpha-1 adrenergic
antagonists. In the presence of an antagonist, the maximum
effect, as well as other pharmacologic parameters in the
concentration-effect curves, may be reduced.1
Figure 2. Contraction induced by phenylephrine in rat thoracic aorta in
the absence or in the presence of pyrimidinone derivatives. Each point
represents mean S.E.M. of different animals. Statistically different from
phenylephrine, p < 0.05 (one wayANOVA, followed by Newmann-Keuls).
Table 3. Contraction induced by phenylephrine in rat thoracic aorta in
the absence or in the presence of the derivatives
In this study, concentration-effect curves to
phenylephrine were constructed in the absence and in
the presence of the derivatives in isolated rat thoracic
aorta. As it can be seen in Figure 2 and Table 3, most of
the derivatives, except 5b, 5g, and 5i, were able to inhibit
the phenylephrine-induced contraction in thoracic aorta
isolated from rats (diminish the maximum effect of the
phenylephrine concentration-effect curve). It was observed
that the presence of electronegative atoms in the phenyl
ring has some influence in improving the biological
activity. The totality of derivatives containing halogen
atoms in their structures showed very good inhibition of the
phenylephrine-induced contraction. These findings show
that electronic effects are important for the maintenance
of the pharmacological activity. In addition, steric effects
seem to rule the activity once 5d (p-ClPh) and 5e (p-BrPh)
showed better percentages of inhibition than 5f (p-FPh).
These results suggest that the molecules may be alpha-1
antagonists. However, more conclusive experiments should
be performed to ensure this assumption.
Contraction of
phenylephrine / g
Percentage of
inhibition / %
Moiety
None
2.68 0.17
0.00 0.00a
1.87 0.24a
2.64 0.36
0.63 0.06a
0.00 0.00a
0.14 0.13a
0.55 0.13a
2.33 0.30
1.34 0.11a
2.31 0.19
0.00 0.00a
-
Prazosin (100 μmol L-1)
5a (100 μmol L-1)
5b (100 μmol L-1)
5c (100 μmol L-1)
5d (100 μmol L-1)
5e (100 μmol L-1)
5f (100 μmol L-1)
5g (100 μmol L-1)
5h (100 μmol L-1)
5i (100 μmol L-1)
5J (100 μmol L-1)
100.0
30.2
1.5
76.5
100.0
94.8
79.5
13.0
50.0
13.8
100.0
Each point represents mean S.E.M. of different animals. aStatistically
different from phenylephrine, p < 0.05 (one way ANOVA, followed by
Newmann-Keuls).

Vol. 28, No. 7, 2017
de Andrade et al.
1273
Conclusions
Brideau, R. J.; Ready, K. A.; Dunn, C. J.; Buxser, S. E.; Int. J.
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In this article we advanced a multicomponent approach
employing ultrasound irradiation under mild conditions
to prepare a series of vasoactive pyrimidinones. This
environmentally friendlier protocol furnishes products
with higher global yields, employs potassium carbonate as
base instead of organic harmful bases and is performed in
2 hours, which is much faster than the usual 2-day route.
Moreover, the molecules were able to inhibit the
phenylephrine-induced contractions in rat thoracic aorta,
suggesting that they act like alpha-1 antagonists. However,
more experiments should be performed to ensure this
assumption.
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Acknowledgments
We thank the Brazilian agencies CNPq and FACEPE
(PRONEX-0788-1.06/08) for financial support.A. N.A. is
grateful to CAPES for a graduate research fellowship. H.
B. W. B. is grateful to CNPq for an undergraduate research
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Submitted: August 11, 2016
9. Vroegop, S. M.; Chapman, D. L.; Decker, D. E.; Galinet, L.A.;
Published online: November 3, 2016