Novel pyrimidinic selenourea induces DNA damage, cell cycle arrest, and apoptosis in human breast carcinoma

Article abstract of DOI:10.1016/j.ejmech.2018.06.026

Novel pyrimidinic selenoureas were synthesized and evaluated against tumour and normal cell lines. Among these, the compound named 3j initially showed relevant cytotoxicity and selectivity for tumour cells. Three analogues of 3j were designed and synthesi

Full text of DOI:10.1016/j.ejmech.2018.06.026

Accepted Manuscript
Novel pyrimidinic selenourea induces DNA damage, cell cycle arrest, and apoptosis
in human breast carcinoma
Flavio A.R. Barbosa, Tâmila Siminski, Rômulo F.S. Canto, Gabriela M. Almeida,
Nádia S.R.S. Mota, Fabiana Ourique, Rozangela Curi Pedrosa, Antonio Luiz Braga
PII:
S0223-5234(18)30517-8
10.1016/j.ejmech.2018.06.026
EJMECH 10496
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 12 December 2017
Revised Date: 7 June 2018
Accepted Date: 9 June 2018
Please cite this article as: F.A.R. Barbosa, Tâ. Siminski, Rô.F.S. Canto, G.M. Almeida, Ná.S.R.S. Mota,
F. Ourique, R.C. Pedrosa, A.L. Braga, Novel pyrimidinic selenourea induces DNA damage, cell cycle
arrest, and apoptosis in human breast carcinoma, European Journal of Medicinal Chemistry (2018), doi:
10.1016/j.ejmech.2018.06.026.
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ACCEPTED MANUSCRIPT

ACCEPTED MANUSCRIPT
Novel pyrimidinic selenourea induces DNA damage, cell cycle arrest, and
apoptosis in human breast carcinoma
a, §
b, §
c
Flavio A. R. Barbosa
, Tâmila Siminski
, Rômulo F. S. Canto , Gabriela M.
b
b
b
b, *
Almeida , Nádia S. R. S. Mota , Fabiana Ourique , Rozangela Curi Pedrosa
,
a, *
Antonio Luiz Braga
a
Laboratório de Síntese de Substâncias de Selênio Bioativas (LabSelen), Departamento
de Química, Universidade Federal de Santa Catarina (UFSC), Florianópolis, SC, Brazil
b
Laboratório de Bioquímica Experimental (LABIOEX), Departamento de Bioquímica,
Universidade Federal de Santa Catarina (UFSC), Florianópolis, SC, Brazil
c
Laboratório de Química Medicinal de Compostos de Selênio (QMCSe), Programa de
pós-graduação em Ciências da Saúde, Universidade Federal de Ciências da Saúde de
Porto Alegre (UFCSPA), Porto Alegre, RS, Brazil.
§
These authors contributed equally to this work
*
Corresponding authors
E-mail address: braga.antonio@ufsc.br
E-mail address: rozangela.pedrosa@ufsc.br
ABSTRACT
Novel pyrimidinic selenoureas were synthesized and evaluated against tumour and
normal cell lines. Among these, the compound named 3j initially showed relevant
cytotoxicity and selectivity for tumour cells. Three analogues of 3j were designed and
synthesized keeping in view the structural requirements of this compound. Almost all

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the tested compounds displayed considerable cytotoxicity. However, 8a, one of the 3j
analogues, was shown to be highly selective and cytotoxic, especially for breast
carcinoma cells (MCF-7) (IC = 3.9 µM). Furthermore, 8a caused DNA damage,
5
0
inhibited cell proliferation, was able to arrest cell cycle in S phase, and induced cell
death by apoptosis in human breast carcinoma cells. Moreover, predictions of
pharmacokinetic properties showed that 8a may present good absorption and
permeation characteristics for oral administration. Overall, the current study established
8
a as a potential drug prototype to be employed as a DNA interactive cytotoxic agent
for the treatment of breast cancer.
Keywords: dihydropyrimidinones, organoselenium, cytotoxicity, DNA damage, cell
cycle arrest, apoptosis, selenoureas
1
.Introduction
Dihydropyrimidinones (DHPMs) are a very important class of six-membered
heterocycles that are obtained by the Biginelli multicomponent reaction [1]. In the field
of medicinal chemistry, DHPMs have been much explored due to the modular nature of
the multicomponent reaction that enables the use of several aldehydes, dicarbonylic
compounds, and ureas, which readily leads to the preparation of a diverse library of
compounds. In addition, these compounds are known for presenting multiple biological
activities, such as antioxidant action [2–4], acetylcholinesterase inhibition [5,6], and
mPGES-1 inhibition [7] among other activities [8]. However, the anticancer activities of
DHPMs are more pronounced, since various reports have described compounds such as
Monastrol and its derivatives as good cytotoxic and antiproliferative agents [9–15].
Molecular hybrids containing the DHPM core are also reported as potential anticancer

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agents, such as DHPM-Coumarin hybrids for breast tumours [16] and DHPM-
semicarbazone hybrids with anticancer activity due to their ability to inhibit human
DNA ligase 1 [17]. More recently, DHPMs containing a fatty acid moiety have been
found to possess antiproliferative activity against C6 glioma cell line [18] and cytotoxic
effects in the melanoma B16F10 cell line [19].
Organoselenium compounds are well known for their antioxidant activity with
the ability to mimic selenoenzyme Glutathione Peroxidase (GPx-like activity) [20,21];
there is also emerging evidence indicating the potential of selenium (Se) compounds in
cancer chemotherapy; these compounds are well known to inhibit cell proliferation and
induce cell death in human cancer cells by apoptosis [22,23]. Se-compounds induce
inhibition of proliferation and promote cellular apoptosis, presumably by decreased
phosphorylation of Akt [24,25] which, through the phosphatidylinositol 3-kinase
PI3K/Akt/mTOR pathway, has been shown to inhibit apoptosis in cancer cells [26] and
promote proliferation and angiogenesis [27]. Furthermore, Se-compounds are related to
apoptotic processes, including cytochrome c-independent caspase-3 activation, down-
regulation of IAP family proteins, and Bax cleavage [28]. Moreover, Suzuki et al. [24],
demonstrated that Se-induced apoptosis in some carcinoma cells is a caspase-dependent
process involving p53 activation. In addition, Se-compounds are involved in the
generation of reactive oxygen species (ROS), DNA damage, mitochondrial dysfunction,
and imbalance of Bcl-2 family expression, which leads to cell death in several human
cancer cells [29].
The pro-oxidant effects of Se-containing compounds have been used as new
tools for developing novel anticancer molecules [30–32]. The insertion of selenium
atom into biologically relevant nuclei such as as indoles [33], naphtoquinones [34,35],
flavonoids [36], and acetylsalicylic acid [37] has been described in the recent years and

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such modification leads to more cytotoxic compounds. The synthesis of molecular
hybrids between ebselen and resveratrol also leads to very potent cytotoxic compounds
with possible mechanisms of action related to inhibition of Thioredoxin Reductase
(
TrxR), another selenoenzyme involved in the redox balance, which leads to cell death
by oxidative stress [38].
Selenoureas, isoselenoureas, and their derivatives have been reported as
antioxidant and anticancer agents used mainly in the design of highly potent molecular
hybrids [39–42]. To the best of our knowledge, the study of pyrimidinic selenoureas has
only two reports in the literature. Klein and co-workers reported the synthesis of a
single seleno-analogue of Monastrol and evaluated its ability to inhibit the Kinesin Eg5
enzyme, however, it caused a two-fold decrease in the activity when compared to
Monastrol [43]. In the second report, Kolb and colleagues evaluated a pyrimidinic
selenourea and its selenazolopyrimidinic derivative as CDC25B phosphatase inhibitors
[
44].
DNA is also an important target for the development of new antitumor drugs,
because interactions between drugs and DNA can result in blockage of cell division and
triggering of cell death [45]. DHPMs and their derivatives have potential to overlap
with DNA base pairs or to bind to the grooves; therefore, they are regarded as potential
new DNA-targeted drugs [46–48]. Organoselenium compounds are also able to cause
DNA damage leading to cell death [49,50].
As part of our research program focused on the synthesis and biological
evaluation of new seleno-DHPMs [51,52], we here report the design, synthesis, and
anticancer evaluation of a series of pyrimidinic selenoureas. These compounds were
designed through molecular hybridization [53] by merging the dihydropyrimidinone
nucleus and the selenourea moiety (Scheme 1) to form one molecule that has the

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pharmacological potential of both classes of compounds.
Scheme 1- Molecular hybridization towards pyrimidinic selenoureas
2
.Results and Discussion
Pyrimidinic selenoureas 3a-k were synthesized via a three-step route (Scheme
), following the previous literature [54], which is based on transforming thioureas into
2
selenoureas using an iso-thiourea intermediate. Thioureas 1a-k were prepared through
the classical Biginelli multicomponent reaction, using different substituted aromatic
aldehydes, ethyl acetoacetate, thiourea and HCl as catalyst [51]. Methylation of
thioureas 1a-k with methyl iodide followed by basic treatment leads to the formation of
iso-thioureas 2a-k with excellent yields. Finally, pyrimidinic selenoureas 3a-k were
obtained by the reaction of iso-thioureas 2a-k with the nucleophilic selenium species
NaSeH in refluxing ethanol for 15 hours. By this strategy, the Se anion adds to the
carbon atom of iso-thiourea, followed by the elimination of methanethiolate as a leaving
group. The compounds were obtained in good to excellent yields (65-99%). Compound
2
d bearing a nitro group was obtained as an aryl amine 3d after reaction with NaSeH,
this reduction was expected since sulphur species such as sodium sulphide have been
used as reducing agents for aromatic nitro compounds [55].

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Scheme 2 - i) HCl, 100 °C, neat, 55-99%; ii) MeI, EtOH, reflux, 1 h, then NaHCO , 72-
3
9
9%; iii) NaSeH, EtOH, reflux, 15h, 65-99%.
Initially, pyrimidinic selenoureas 3a-k were evaluated against two cancer cell
lines, MCF-7 and HeLa, and the normal cell line McCoy. The cells were treated for
different periods of time with different concentrations of the pyrimidinic selenoureas
3
a-k followed by screening for cell cytotoxicity by MTT assay. Table 1 shows values
obtained from IC (half-maximal inhibitory concentration) for compounds 3a-k and
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two standards, Monastrol and the anticancer drug Fluorouracil (5-FU), 72 h post-
treatment. As previously demonstrated, both DHPMs and Se-compounds induced time-
dependent cytotoxicity and growth inhibition in different cancer cell lines [14,24].

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Table 1 - IC values of the pyrimidinic selenoureas 3a-k in different cancer cell lines
5
0
and in normal cells.
IC (µM)
5
0
Compound
MCF-7
452.2
92.9
HeLa
909.1
205.1
390.2
512.3
283.8
254.3
258.3
149.1
152.6
87.6
McCoy
251.8
175.9
252.7
289.1
212.9
199.1
107.9
135.6
143.9
57.7
3
a
3
b
3
c
175.8
207.1
200.6
301
3
d
3
e
3
f
3
g
169.4
247.8
251.7
25.2
3
h
3
i
3
j
3
k
181.1
59.1
224.3
176.9
7.00
194
Monastrol
-FU
26.8
5
63.0
15.7
Cells were treated for 72 h with compounds (0.1 – 1000 µM) and the cytotoxicity data
were obtained by MTT assay. Data from three independent experiments.
As seen in Table 1, most of the compounds exhibited high IC values for both
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0
cancer cell lines, however, compound 3j presented the lowest IC value among other
5
0
pyrimidinic selenoureas, with IC of 25.2 µM for MCF-7 cells and 87.6 µM for HeLa,
5
0
more than two times the activity of the standard Monastrol. Also, compound 3j was 2,5
times more efficient than the 5-FU for MCF-7 cells.

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Based on these results, we designed three new analogues of compound 3j,
keeping the important naphthyl moiety in the structure, aiming for an enhancement in
antitumor activity. Two main modifications were planned for its structure: modification
of the selenourea moiety in 3j by methylation of the selenium atom, leading to iso-
selenourea 4a, and lateral chain cyclization of the heterocycle, leading to compounds
8
a-b (Scheme 3). The latter modifications are well known for the structure-activity
relationships in DHPM cores as antitumor agents, and these modifications usually lead
to more active compounds [56].
Scheme 3 -Design of 3j analogues by methylation of selenium and lateral chain
cyclization
Methylation of the selenium of 3j occurred successfully applying the same
methodology usually used for sulphur analogues, affording the product 4a in 98% yield
(Scheme 4).

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Scheme 4 - iv) MeI, EtOH, reflux, 1 h, then NaHCO , 98%
3
Two other analogues, 8a-b, were synthesized using the same strategy applied for
selenoureas 3a-k, replacing the ethyl acetoacetate by cyclohexanedione and dimedone.
The desired compounds were obtained in good to excellent yields, except for 6b, which
was obtained in a moderate yield (Scheme 5).
O
O
O
H
O
O
v
vi
vii
O
NH
N
NH
Se
H N
2
R''
R''
N
S
R''
R''
N
S
R''
R''
N
H
H
H
H N
S
R''
R''
2
R'' = CH : 6a
R'' = CH : 7a
R'' = CH : 8a
3
3
3
R'' = H: 6b
R'' = H: 7b
R'' = H: 8b
Scheme 5 - v) HCl, 100°C, neat, 66-35%; vi) MeI, EtOH, reflux, 1 h, then NaHCO , 90-
3
9
9%; vii) NaSeH, EtOH, reflux, 15 h, 92-86%.
With the three new analogues 4a and 8a-b in hand, it was possible to evaluate
their activity against MCF-7 and HeLa cells; the results for their IC50 values are
presented in Table 2. In general, the three modifications resulted in more cytotoxic
compounds when compared to 3j. Compound 4a containing an isoselenourea showed an
IC value lower than 3j. However, 4a also showed a decrease in selectivity, since it was
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0
very toxic to normal cells. According Badisa et al. [57], selectivity index (SI)
demonstrates the differential activity of a compound for a tumour cell, and a SI value

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greater than two indicate low toxicity for normal cells and high toxicity against tumour
cell lines.
The compounds presenting the cyclized lateral chain also lead to more cytotoxic
compounds (8a and 8b). Compound 8a was particularly interesting because the
presence of a dimethyl group in the lateral chain caused a significant increase in
cytotoxicity, leading to a decrease in viability of tumour cells, with IC values showing
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0
activities approximately 15 times more pronounced than Monastrol and 16 times more
citotoxic when compared to the drug 5-FU for MCF-7 cells. In addition, these
compounds showed substantial activity without further damage to normal cells (Table
2
). To our delight, 8a presented higher selectivity than the standards monastrol and 5-
FU, showing its great potential for further studies. This structural alteration and
potentiation of the antitumor effect for 8a is in accord with that proposed by Müller et
al. [58], who reported that enastron and dimethylenastron, both cyclized analogues of
Monastrol, increased cytotoxic effects against human glioblastoma cells.
Table 2 - IC and SI values of the pyrimidinicselenoureas4a, 8a-b and 3j in different
5
0
cancer cell lines and in normal cells.
IC (µM)
Selectivity Index (SI)
5
0
Compound
MCF-7
HeLa
McCoy
MCF-7
HeLa
4
8
8
a
a
b
8.8
11.2
0.01
0.001
0.0009
3.9
12.2
26.2
87.6
176.9
7.00
425.8
22.4
57.7
26.8
15.7
109.1
1.7
34.9
0.8
0.6
0.2
2.2
13.1
25.2
59.1
63.0
3
j
2.2
Monastrol
-FU
0.4
5
0.2

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SI = IC of compound in normal cell line/IC of the compound in cancer cell line. Data
5
0
50
from the IC mean of three independent experiments.
5
0
As seen that 8a presented the most relevant results in terms of viability and
selectivity, other biological assays were conducted in human breast carcinoma in order
to elucidate the mechanisms of such effects.
The antiproliferative effect of 8a (2.4 µM) against MCF-7 cells was evaluated
over 72 h of treatment and the results are presented in Fig. 2A. The results indicated that
8
a inhibited cell proliferation, as shown by the reduction in the number of colonies
(40%) in comparison to non-treated cells. Guido et al. [59] reported that DHPMs have
been associated with inhibition of proliferation and angiogenesis. These effects may be
related to kinesin Eg5 [60], a protein involved in the formation and function of the
mitotic spindle, and to DNA damage [61]. Compound 8a was also able to induce MCF-
7
cell death (Fig. 2B) by apoptosis and necrosis. Apoptosis, the most pursued goal in
anticancer drug development [62], was significantly increased after treatment with
compound 8a (p < 0.001). This result is very interesting since MCF-7 cells may be
resistant to apoptosis due to the possible absence of caspase-3 [63].

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Figure 2. Antiproliferative effect (A) and induction of cell death (B) after treatment of
MCF-7 with non-toxic concentrations of compound 8a (2.4 µM) for 72 h. Data were
obtained from three independent experiments. (***) denotes statistical differences at p <
0
.001 compared to control cells (non-treated).
DNA is also a target for developing new potential drugs [64], and we
investigated whether 8a is able to interact with DNA. Figure 3A and 3B show data
concerning the absorption scanning of CT-DNA alone and after treatment with
compound 8a. Electronic absorption spectroscopy is an effective method to examine the
binding modes of drug with DNA. Hypochromism was observed when 8a, in increasing
concentrations, was placed in the presence of CT-DNA (Fig. 3A). This effect results

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from contraction and changes in the conformation of DNA [65]. The possible DNA
intercalation effect of 8a was investigated (Fig. 3B). For this, propidium iodide was
used because it displays an enhancement of DNA fluorescence efficiency due to its
strong intercalation between adjacent base pairs. 8a was able to remove propidium
iodide and insert between the DNA strands, decreasing CT-DNA fluorescence, which is
indicative of intercalation. Our results are in agreement with those obtained by Wang et
al. [46] who showed that the potential antitumor effects of dihydropyrimidinone
derivatives are related to its DNA binding properties.
Next, the ability of compound 8a to cause DNA fragmentation in MCF-7 cells
was evaluated by Comet assay and the results are shown in Figure 3C. After 72 h of
treatment we observed a significant increase in the total DNA damage caused by 8a (p <
0
.001) in MCF-7 cells treated with non-toxic concentrations. Double-stranded DNA
breaks have been shown to correlate with DNA-targeting chemotherapeutics leading to
inhibition of proliferation and cell death [64]. Planar and non-planar, aromatic and
heterocyclic compounds, such as dihydropyrimidinone derivatives have potential for
overlap with DNA base pairs or binding to the grooves [46]. Also, Se-compounds may
be involved in induction of DNA breakdown caused by generation of intracellular ROS
[
66]. Compounds that cause DNA damage and consequent cell cycle arrest have been
considered promising agents for chemotherapy [67].

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Figure 3. The interaction (A) and intercalating ability via fluorescence intensity of
propidium iodide (300 µM) and doxorubicin as a positive control for intercalation (B) of
compound 8a (0-350 µM) with CT-DNA (150 µM) were evaluated. MCF-7 cells were
treated with 8a (2.4 µM) for 72 h. DNA damage index was determined by Comet assay
(C) and DNA fragmentation scores are shown in the box below the graph. (***) denotes
statistically significant differences at p < 0.001 compared to control cells (non-treated).

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The assessment of the cell cycle of cells treated with 8a was realized and Figure
4
A shows distribution of MCF-7 cells in different phases of the cell cycle. The
treatment with 8a leads to an increase in the sub-G1 area of 15.5% (indicative of
apoptosis). Additionally, the number of cells was significantly reduced in S phase by up
to 4.9%, as shown in Figure 4B. Therefore, 8a showed a large safety to normal cells,
and the cytotoxic effects on breast carcinoma cells are related to DNA damage and cell
cycle arrest, which leads to cell death by apoptosis.
Figure 4. Changes in MCF-7 cell cycle induced by treatment were measured (A) and
percentage of cell distribution in cap S was evaluated (B). Data were obtained from
three independent experiments. (***) denotes statistically significant differences at p <
0
.001 compared to control cells (non-treated).

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Moreover, the pharmacokinetic properties for compound 8a were predicted
using the open-source software Molinspiration [68] and ChemBioDraw Ultra 11.0.
According to Lipinski’s rule of 5 [69] poor absorption or permeation will be more likely
if a drug candidate presents molecular weight > 500, number of hydrogen bond donors
>
5, number of hydrogen bond acceptors > 10 and Clog P > 5.0. Another two relevant
properties in this type of analysis were introduced by Veber and co-workers [70], and
these state that a drug prototype should not have more than 10 rotatable bonds and its
2
topological polar surface area should be ≤ 140 Å .
As seen in Table 3, compound 8a satisfied such criteria for presenting
potentially- good absorption and permeation after oral administration, which enhance its
potential as a drug prototype.
Table 3. Predictions of pharmacokinetic properties for compound 8a.
Property
CLogP
Value
3.58
Molecular Weight
383.35
a
2
tPSA (Å )
41.12
b
HBAc
3
2
1
HBD
d
NRB
a
b
c
tPSA, Topological polar surface area; HBA, H-bond acceptors; HBD, H-bond
donors; NRB, Number of rotatable bonds.
d

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3
. Conclusion
DNA damage, cell cycle arrest, and induction of cell death by apoptosis are
attractive strategies to selectively kill cancer cells. In this work, we demonstrated the
synthesis of novel pyrimidinic selenoureas and their cytotoxicity against tumour cell
lines. Among the compounds evaluated, 8a appears to be promising for the
development of an antitumor agent. This compound caused DNA damage, cell cycle
arrest, antiproliferative effects and cell death by apoptosis. Compound 8a also showed
remarkable selective cytotoxicity for tumour cells, being more selective than the drug 5-
FU, mainly for human breast carcinoma. The prediction of pharmacokinetic properties
indicated that 8a may present good absorption and permeation characteristics. Hence,
compound 8a is a potential drug prototype for the treatment of breast cancer. More in-
depth studies are currently being conducted in our laboratories to clarify and offer more
molecular details concerning the mechanisms of action of compound 8a.
4
.Materials and Methods
4
.1. Chemicals
1
13
NMR spectra ( H NMR and C NMR) were recorded on a Varian AS-400 or
Bruker Avance 200 spectrometer. Chemical shifts (δ) are reported (in ppm) relative to
1
13
the TMS ( H NMR) and the solvent ( C NMR). APPI or APCI-micrOTOF-Q II
measurements were performed with a micrOTOF Q-II (Bruker Daltonics) mass
spectrometer equipped with an automatic syringe pump (KD Scientific) for sample
injection. The mass spectrometer was operated in the positive ion mode. The sample
was injected using a constant flow (3 µL/min). The solvent was a chloroform/methanol
mixture. The APPI-micrOTOF-Q II instrument was calibrated in the mass range of 50–
3
000 m/z using an internal calibration standard (low concentration tuning mix solution)

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supplied by Agilent Technologies. Data were processed employing Bruker Compass
Data Analysis software (version 4.0). Thin layer chromatography (TLC) was conducted
using Merck Silica Gel GF254 (0.25 mm thickness). For visualization, the TLC plates
were either placed under ultraviolet light or stained with iodine vapour or acidic
vanillin. Melting points were determined using a microscope coverslip on a Micro
Chemical MQA PF digital apparatus and are uncorrected. Dulbecco's modified Eagle’s
medium (DMEM), foetal bovine serum (FBS) and antibiotics were purchased from
Gibco (USA). The following material was purchased from Sigma-Aldrich: calf thymus
DNA (CT-DNA; Cat. D4522), agarose (Cat. A6013), dimethyl sulphoxide (DMSO; Cat.
D8418), bovine serum albumin (BSA; Cat. A2153), propidium iodide (Cat. P4170),
acridine orange (Cat. A6014), nocodazole (Cat. M1404) and 3-(4,5-dimethyl-2-
thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT; Cat. M5655). All common
reagents and solvents were used as purchased unless otherwise noted.
4
4
.2.Synthetic procedures
.2.1. General procedure for the synthesis of compounds 1a-k and 6a-b:
The procedure described in the literature [51] was followed. To a two-necked
round-bottom flask, the appropriated aldehyde (10 mmol), ethyl acetoacetate or
dimedone or cyclohexanedione (10 mmol), thiourea (20 mmol) and 5 drops of
concentrated HCl were added. The reaction mixture was stirred at 100°C for the time
required for consumption of the starting materials, which was verified by TLC. After
this time, the reaction mixture was poured into crushed ice and water. The precipitate
was filtered and dried. The compounds were used in the next step without further
purification.

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4
.2.1.1.
Ethyl
6-methyl-4-phenyl-2-thioxo-1,2,3,4-tetrahydropyrimidine-5-
carboxylate (1a)
1
White solid, m.p. 204-207 ºC [Lit. 210ºC][44], 72% yield; H NMR (400 MHz, DMSO-
d6) δ 10.35 (s, 1H), 9.67 (s, 1H), 7.34 (t, J = 7.3 Hz, 2H), 7.28 – 7.22 (m, 3H), 5.19 (d, J
1
3
=
3.6 Hz, 1H), 4.00 (q, J = 6.9 Hz, 2H), 2.30 (s, 3H), 1.09 (t, J = 7.1 Hz, 3H). C NMR
(
101 MHz, DMSO-d ) δ 174.30, 165.19, 145.10, 143.57, 128.62, 127.75, 126.47,
6
1
00.79, 59.67, 54.14, 17.25, 14.07.
4
.2.1.2.
Ethyl
4-(3-hydroxyphenyl)-6-methyl-2-thioxo-1,2,3,4-
tetrahydropyrimidine-5-carboxylate (1b)
1
White solid, m.p. 150-153 ºC [Lit. 152-154ºC][44], 70% yield; H NMR (200 MHz,
DMSO- d ) δ 10.30 (s, 1H), 9.61 (s, 1H), 9.46 (bs, 1H), 7.12 (t, J = 7.8 Hz, 1H), 6.66 (d,
6
J = 6.5 Hz, 3H), 5.10 (d, J = 2.6 Hz, 1H), 4.01 (q, J = 6.8 Hz, 2H), 2.28 (s, 3H), 1.11 (t,
13
J = 6.9 Hz, 3H). C NMR (101 MHz, DMSO- d ) δ 174.25, 165.28, 157.54, 144.89,
6
1
29.59, 117.11, 114.72, 113.34, 100.89, 59.72, 54.05, 17.27, 14.13.
4
.2.1.3.
Ethyl
4-(4-hydroxy-3-methoxyphenyl)-6-methyl-2-thioxo-1,2,3,4-
tetrahydropyrimidine-5-carboxylate (1c)
1
White solid, m.p. 231-233 ºC [Lit. 238-239ºC][12], 55% yield; H NMR (400 MHz,
DMSO- d ) δ 10.26 (s, 1H), 9.57 (s, 1H), 9.03 (bs, 1H), 6.79 (d, J = 1.9 Hz, 1H), 6.73
6
(d, J = 8.1 Hz, 1H), 6.59 (dd, J = 8.1, 1.9 Hz, 1H), 5.09 (d, J = 3.5 Hz, 1H), 4.01 (q, J =
1
3
7
.0 Hz, 2H), 3.73 (s, 3H), 2.28 (s, 3H), 1.11 (t, J = 7.1 Hz, 3H). C NMR (101 MHz,
DMSO- d ) δ 174.09, 165.34, 147.41, 146.22, 144.70, 134.65, 118.62, 115.48, 110.98,
6
1
01.07, 59.64, 55.64, 53.76, 17.22, 14.17.

ACCEPTED MANUSCRIPT
4
5
.2.1.4.
Ethyl 6-methyl-4-(3-nitrophenyl)-2-thioxo-1,2,3,4-tetrahydropyrimidine-
-carboxylate (1d)
1
White solid, m.p. 200-205 ºC [Lit. 206-209ºC][71], 65% yield; H NMR (400 MHz,
DMSO- d ) δ 10.52 (s, 1H), 9.78 (s, 1H), 8.17 – 8.12 (m, 1H), 8.08 (s, 1H), 7.67 (d, J =
6
5
.3 Hz, 2H), 5.34 (d, J = 3.6 Hz, 1H), 4.07 – 3.95 (m, 2H), 2.31 (s, 3H), 1.09 (t, J = 7.1
1
3
Hz, 3H). C NMR (101 MHz, DMSO- d ) δ 174.56, 164.91, 147.86, 146.09, 145.56,
6
1
33.10, 130.47, 122.78, 121.25, 99.89, 59.86, 53.58, 17.31, 14.00.
4
.2.1.5.
Ethyl
4-(benzo[d][1,3]dioxol-5-yl)-6-methyl-2-thioxo-1,2,3,4-
tetrahydropyrimidine-5-carboxylate (1e)
1
Yellowish solid, m.p. 156-159 ºC [Lit. 175ºC][44], 70% yield; H NMR (200 MHz,
DMSO- d ) δ 10.32 (s, 1H), 9.61 (s, 1H), 6.87 (d, J = 7.9 Hz, 1H), 6.72 – 6.65 (m, 2H),
6
5
7
1
.99 (s, 2H), 5.09 (d, J = 3.5 Hz, 1H), 4.01 (q, J = 7.0 Hz, 2H), 2.29 (s, 3H), 1.11 (t, J =
1
3
.1 Hz, 3H). C NMR (101 MHz, DMSO- d ) δ 174.05, 165.14, 147.41, 146.73,
6
45.04, 137.48, 119.67, 108.18, 106.75, 101.11, 100.73, 59.62, 53.72, 17.20, 14.07.
4
.2.1.6.
Ethyl
4-(4-methoxyphenyl)-6-methyl-2-thioxo-1,2,3,4-
tetrahydropyrimidine-5-carboxylate (1f)
1
Beige solid, m.p. 151-154 ºC [Lit. 150-151ºC][72], 99% yield; H NMR (200 MHz,
DMSO- d ) δ 10.29 (s, 1H), 9.60 (s, 1H), 7.13 (d, J = 8.5 Hz, 2H), 6.89 (d, J = 8.6 Hz,
6
2
H), 5.12 (d, J = 3.5 Hz, 1H), 4.00 (q, J = 7.0 Hz, 2H), 3.72 (s, 3H), 2.29 (s, 3H), 1.10
1
3
(
t, J = 7.0 Hz, 3H). C NMR (101 MHz, DMSO- d ) δ 174.04, 165.20, 158.77, 144.79,
6
1
35.74, 127.66, 113.91, 101.00, 59.60, 55.13, 53.49, 17.19, 14.08.

ACCEPTED MANUSCRIPT
4
.2.1.7.
Ethyl
6-methyl-2-thioxo-4-p-tolyl-1,2,3,4-tetrahydropyrimidine-5-
carboxylate (1g)
1
Beige solid, m.p. 180-182 ºC [Lit. 185-188ºC][73], 85% yield; H NMR (400 MHz,
DMSO- d ) δ 10.31 (s, 1H), 9.63 (s, 1H), 7.14 (d, J = 8.2 Hz, 2H), 7.11 (d, J = 8.3 Hz,
6
2
H), 5.15 (d, J = 3.6 Hz, 1H), 4.00 (q, J = 7.1 Hz, 2H), 2.29 (s, 3H), 2.25 (s, 3H), 1.10
1
3
(
t, J = 7.1 Hz, 3H). C NMR (101 MHz, DMSO- d ) δ 174.21, 165.20, 144.93, 140.66,
6
1
36.96, 129.11, 126.36, 100.88, 59.62, 53.81, 20.72, 17.21, 14.08.
4
.2.1.8.
Ethyl
4-(3,4-dimethoxyphenyl)-6-methyl-2-thioxo-1,2,3,4-
tetrahydropyrimidine-5-carboxylate (1h)
1
Orange solid, m.p. 171-174 ºC [Lit. 178ºC][44], 72% yield; H NMR (400 MHz,
DMSO- d ) δ 10.32 (s, 1H), 9.62 (s, 1H), 6.91 (d, J = 8.3 Hz, 1H), 6.84 (d, J = 1.8 Hz,
6
1
H), 6.71 (dd, J = 8.3, 1.9 Hz, 1H), 5.13 (d, J = 3.6 Hz, 1H), 4.02 (q, J = 7.1 Hz, 3H),
1
3
3
.72 (s, 3H), 3.71 (s, 3H), 2.29 (s, 3H), 1.12 (t, J = 7.1 Hz, 3H). C NMR (101 MHz,
DMSO- d ) δ 174.22, 165.28, 148.57, 148.38, 144.92, 136.02, 118.21, 111.83, 110.48,
6
1
00.88, 59.66, 55.56, 55.46, 53.65, 17.21, 14.15.
4
.2.1.9.
Ethyl
4-(3-bromophenyl)-6-methyl-2-thioxo-1,2,3,4-
tetrahydropyrimidine-5-carboxylate (1i)
1
White solid, m.p. 165-168 ºC [Lit. 171ºC][74], 71% yield; H NMR (400 MHz, DMSO-
d6) δ 10.43 (s, 1H), 9.69 (s, 1H), 7.48 (d, J = 7.9 Hz, 1H), 7.38 (s, 1H), 7.32 (t, J = 7.8
Hz, 1H), 7.22 (d, J = 7.8 Hz, 1H), 5.17 (s, 1H), 4.08 – 3.95 (m, 2H), 2.30 (s, 3H), 1.10
1
3
(
t, J = 7.1 Hz, 4H). C NMR (101 MHz, DMSO- d ) δ 174.38, 164.99, 146.07, 145.63,
6
1
31.01, 130.62, 129.33, 125.46, 121.74, 100.15, 59.77, 53.64, 17.28, 14.04.

ACCEPTED MANUSCRIPT
4
.2.1.10.
Ethyl
6-methyl-4-(naphthalen-2-yl)-2-thioxo-1,2,3,4-
tetrahydropyrimidine-5-carboxylate (1j)
1
White solid, m.p. 170-174 ºC [Lit. 177ºC][44], 76% yield; H NMR (200 MHz, DMSO-
d6) δ 10.41 (s, 1H), 9.77 (s, 1H), 7.93 – 7.89 (m, 3H), 7.69 (s, 1H), 7.52 – 7.46 (m, 3H),
1
3
5
.37 (s, 1H), 4.00 (q, J = 7.0 Hz, 2H), 2.35 (s, 3H), 1.08 (t, J = 7.0 Hz, 3H). C NMR
(
50 MHz, DMSO- d ) δ 174.24, 165.15, 145.27, 140.81, 132.65, 132.46, 128.53,
6
1
27.92, 127.52, 126.43, 126.16, 124.97, 124.82, 100.51, 59.60, 54.39, 17.27, 14.04.
4
.2.1.11.
Ethyl
4-(4-hydroxyphenyl)-6-methyl-2-thioxo-1,2,3,4-
tetrahydropyrimidine-5-carboxylate (1k)
1
Yellowish solid, m.p. 202-206 ºC [Lit. 202ºC][44], 73% yield; H NMR (400 MHz,
DMSO- d ) δ 10.25 (s, 1H), 9.56 (s, 1H), 9.46 (s, 1H), 7.02 (d, J = 8.4 Hz, 2H), 6.71 (d,
6
J = 8.4 Hz, 2H), 5.07 (d, J = 3.3 Hz, 1H), 3.99 (q, J = 6.9 Hz, 2H), 2.28 (s, 3H), 1.09 (t,
13
J = 7.1 Hz, 3H). C NMR (101 MHz, DMSO- d ) δ 173.95, 165.32, 157.01, 144.63,
6
1
34.23, 127.79, 115.26, 101.22, 59.64, 53.69, 17.25, 14.14.
4
.2.1.12.
7,7-dimethyl-4-(naphthalen-2-yl)-2-thioxo-1,2,3,4,7,8-
hexahydroquinazolin-5(6H)-one (6a)
1
White solid, m.p. above 250 ºC, 66% yield. H NMR (200 MHz, DMSO-d ) δ 10.65 (s,
6
1
8
=
H), 9.80 (s, 1H), 7.92 – 7.88 (m, 3H), 7.70 (s, 1H), 7.53 – 7.48 (m, 2H), 7.40 (d, J =
.3 Hz, 1H), 5.37 (d, J = 2.7 Hz, 1H), 2.45 (s, 2H), 2.25 (d, J = 16.2 Hz, 1H), 2.06 (d, J
1
3
16.1 Hz, 1H), 1.03 (s, 3H), 0.89 (s, 3H). C NMR (50 MHz, DMSO-d ) δ 193.66,
6
1
1
3
74.63, 148.81, 140.67, 132.61, 132.35, 128.38, 127.84, 127.48, 126.38, 126.08,
-1
25.01, 124.68, 107.93, 52.54, 49.85, 38.55, 32.23, 28.77, 26.72. IR (KBr), (ν, cm ):
530, 3451, 3249, 3204, 3008, 2957, 2927, 2888, 1668, 1646, 1623, 1601, 1575, 1466,

ACCEPTED MANUSCRIPT
1
3
373. HRMS (APPI) m/z calculated for C H N OS [M+H] 337.1369; found
2
0
20
2
37.1368.
4
.2.1.13.
4-(naphthalen-2-yl)-2-thioxo-1,2,3,4,7,8-hexahydroquinazolin-5(6H)-one
(6b)
1
Off white solid, m.p. above 250 ºC, 35% yield. H NMR (200 MHz, DMSO-d ) δ 10.66
6
(s, 1H), 9.78 (s, 1H), 7.92 – 7.87 (m, 3H), 7.68 (s, 1H), 7.53 – 7.48 (m, 2H), 7.42 (d, J =
8
1
1
2
2
3
.4 Hz, 1H), 5.37 (d, J = 3.0 Hz, 1H), 2.51 – 2.50 (m, 2H), 2.32 – 2.25 (m, 2H), 2.03 –
1
3
.77 (m, 2H). C NMR (50 MHz, DMSO-d ) δ 193.93, 174.48, 150.85, 140.57,
6
32.60, 132.34, 128.35, 127.85, 127.42, 126.29, 126.03, 124.79, 108.82, 52.18, 36.32,
-1
5.34, 20.49. IR (KBr), (ν, cm ): 3261, 3212, 3182, 3106, 3055, 3021, 2958, 2886,
860, 1622, 1568, 1507, 1458. HRMS (APPI) m/z calculated for C H N OS [M+H]
1
8
16
2
09.1056; found 309.1052.
4
.2.2.
General procedure for the synthesis of isothioureas2a-k, 7a-b and
isoselenourea4a
The procedures described previously in the literature [75] were followed. In a
two-necked round-bottom flask, the appropriate thioureas 1a-k, 6a-b or selenourea 3j (5
mmol) were solubilized in ethanol, followed by the addition of methyl iodide (6 mmol).
The reaction mixture was stirred at reflux temperature for the time required for
consumption of the starting material, which was verified by TLC. After this time, the
reaction mixture was treated with 20 ml of a NaHCO (Sat.) solution. The reaction
3
mixture was extracted with ethyl acetate/water and the organic phase dried over MgSO4

ACCEPTED MANUSCRIPT
and concentrated under vacuum. The resulting products presented high purity and were
used in the next step without further purification.
4
.2.2.1.
Ethyl
6-methyl-2-(methylthio)-4-phenyl-1,4-dihydropyrimidine-5-
carboxylate (2a)
1
White solid, m.p. 172-174 ºC [Lit. 171-172ºC][76], 99% yield; H NMR (200 MHz,
DMSO- d ) δ 9.59 (s, 1H), 7.32 – 7.20 (m, 5H), 5.49 (s, 1H), 4.00 (q, J = 7.0 Hz, 2H),
6
1
3
2
.30 (s, 3H), 2.24 (s, 3H), 1.11 (t, J = 7.1 Hz, 3H). C NMR (50 MHz, DMSO- d ) δ
6
1
66.21, 145.46, 128.15, 126.76, 126.43, 58.98, 14.07, 12.52.
4
.2.2.2.
Ethyl
4-(3-hydroxyphenyl)-6-methyl-2-(methylthio)-1,4-
dihydropyrimidine-5-carboxylate (2b)
1
Beige solid, m.p. 201-204ºC, 91% yield; H NMR (400 MHz, DMSO- d ) δ 9.59 (s,
6
1
H), 9.28 (s, 1H), 7.05 (t, J = 7.1 Hz, 1H), 6.66 (s, 1H), 6.63 (d, J = 1.4 Hz, 1H), 6.58
(d, J = 6.2 Hz, 1H), 5.43 (s, 1H), 4.01 (q, J = 7.0 Hz, 2H), 2.29 (s, 3H), 2.22 (s, 3H),
1
3
1
1
2
.13 (t, J = 7.1 Hz, 3H). C NMR (50 MHz, DMSO- d ) δ 166.37, 157.28, 145.77,
6
-1
29.09, 117.22, 113.48, 59.09, 14.20, 12.63. IR (KBr) (ν, cm ): 3290, 3192, 3070,
978, 2931, 2811, 2713, 2592, 2468, 1664, 1591, 1485, 1452. HRMS (APPI) m/z
calculated for C H N O S 307.1111 [M+H]; found 307.1116.
1
5
18
2
3
4
.2.2.3.
dihydropyrimidine-5-carboxylate (2c)
Beige solid, m.p. 163-164ºC, 87% yield; H NMR (400 MHz, DMSO- d ) δ 9.54 (s,
Ethyl
4-(4-hydroxy-3-methoxyphenyl)-6-methyl-2-(methylthio)-1,4-
1
6
1
H), 8.83 (s, 1H), 6.78 (s, 1H), 6.68 (d, J = 8.0 Hz, 1H), 6.58 (d, J = 7.3 Hz, 1H), 5.41

ACCEPTED MANUSCRIPT
(s, 1H), 4.04 – 3.98 (m, 3H), 3.71 (s, 3H), 2.30 (s, 3H), 2.22 (s, 3H), 1.13 (t, J = 7.1 Hz,
1
3
3
H). C NMR (101 MHz, DMSO- d ) δ 166.40, 147.17, 118.73, 115.25, 111.07, 58.96,
6
-1
5
5.60, 14.17, 12.58. IR (KBr) (ν, cm ): 3304, 2994, 2930, 2831, 2737, 2592, 1658,
1
611, 1595, 1519. HRMS (APPI) m/z calculated for C H N O S 337.1217 [M+H];
1
6
20
2
4
found 337.1217.
4
5
.2.2.4.
Ethyl 6-methyl-2-(methylthio)-4-(3-nitrophenyl)-1,4-dihydropyrimidine-
-carboxylate (2d)
1
White solid, m.p. 215-218 ºC [Lit. 220-221ºC][77], 99% yield; H NMR (400 MHz,
CDCl ) δ 8.17 (s, 1H), 8.08 (dd, J = 8.1, 2.3 Hz, 1H), 7.69 (dd, J = 7.7, 1.1 Hz, 1H),
3
7
.49 – 7.45 (m, 1H), 5.78 (s, 1H), 4.18 – 4.10 (m, 2H), 2.40 (s, 3H), 2.34 (s, 3H), 1.23
1
3
(
t, J = 7.1 Hz, 3H). C NMR (101 MHz, CDCl ) δ 166.60, 148.28, 147.05, 133.42,
3
1
29.26, 122.12, 122.00, 60.15, 14.24, 13.48.
4
.2.2.5.
Ethyl
4-(benzo[d][1,3]dioxol-5-yl)-6-methyl-2-(methylthio)-1,4-
dihydropyrimidine-5-carboxylate (2e)
Yellow solid, m.p. 118-121 ºC, 95% yield; H NMR (200 MHz, CDCl ) δ 6.82 (s, 1H),
1
3
6
3
1
.72 (t, J = 8.5 Hz, 2H), 5.90 (s, 2H), 5.53 (s, 1H), 4.10 (q, J = 7.1 Hz, 2H), 2.40 (s,
1
3
H), 2.31 (s, 3H), 1.20 (t, J = 7.1 Hz, 3H). C NMR (101 MHz, CDCl ) δ 190.59,
3
66.98, 147.61, 146.63, 139.19, 120.13, 107.93, 107.60, 100.91, 59.87, 14.29, 13.47. IR
-1
(
KBr) (ν, cm ): 3325, 3278, 3223, 3078, 2978, 2931, 2906, 2782, 1854, 1668, 1617,
1
3
501, 1485. HRMS (APCI) m/z calculated for C H N O S [M+H] 335.1060; found
16 18 2 4
35.1059.

ACCEPTED MANUSCRIPT
4
.2.2.6.
Ethyl
4-(4-methoxyphenyl)-6-methyl-2-(methylthio)-1,4-
dihydropyrimidine-5-carboxylate (2f)
1
Yellowish solid, m.p. 126-129 ºC [Lit. 135-136ºC][77], 91% yield; H NMR (400
MHz, CDCl ) δ 7.23 (d, J = 8.5 Hz, 2H), 6.81 (d, J = 8.6 Hz, 2H), 5.55 (s, 1H), 4.09 (q,
3
13
J = 7.1 Hz, 2H), 3.77 (s, 3H), 2.41 (s, 3H), 2.32 (s, 3H), 1.20 (t, J = 7.1 Hz, 3H). C
NMR (101 MHz, CDCl ) δ 167.00, 158.96, 137.47, 128.14, 113.86, 59.85, 55.34,
3
1
4.36, 13.59.
4
.2.2.7.
Ethyl
6-methyl-2-(methylthio)-4-p-tolyl-1,4-dihydropyrimidine-5-
carboxylate (2g)
1
Beige solid, m.p. 169-171 ºC [Lit. 175-176ºC][77], 99% yield; H NMR (400 MHz,
CDCl ) δ 7.22 (d, J = 8.1 Hz, 2H), 7.10 (d, J = 7.9 Hz, 2H), 5.61 (s, 1H), 4.10 (q, J =
3
1
3
7
.1 Hz, 2H), 2.44 (s, 3H), 2.34 (s, 3H), 2.32 (s, 3H), 1.21 (t, J = 7.1 Hz, 3H). C NMR
(
101 MHz, CDCl ) δ 166.88, 141.90, 137.05, 129.47, 129.18, 128.46, 126.92, 126.61,
3
5
9.90, 21.20, 14.34, 13.80, 13.65.
4
.2.2.8.
Ethyl
4-(3,4-dimethoxyphenyl)-6-methyl-2-(methylthio)-1,4-
dihydropyrimidine-5-carboxylate (2h)
Orange solid, m.p. 58-61 ºC, 99% yield; H NMR (400 MHz, CDCl ) δ 6.91 (s, 1H),
1
3
6
.84 (dd, J = 8.3, 1.8 Hz, 1H), 6.77 (d, J = 8.3 Hz, 1H), 5.58 (s, 1H), 4.11 (q, J = 7.1 Hz,
1
3
2
H), 3.84 (s, 6H), 2.42 (s, 3H), 2.33 (s, 3H), 1.21 (t, J = 7.1 Hz, 3H). C NMR (101
MHz, CDCl ) δ 167.02, 148.88, 148.33, 137.87, 118.94, 111.21, 110.69, 59.83, 55.93,
3
-1
1
4.36, 13.55. IR (KBr) (ν, cm ): 3594, 3427, 3217, 3078, 2986, 2962, 2929, 2904,
2
833, 2042, 1850, 1648, 1593, 1513, 1477. HRMS (APCI) m/z calculated for
C H N O S [M+H] 351.1373; found 351.1375.
17
22
2
4

ACCEPTED MANUSCRIPT
4
.2.2.9.
Ethyl
4-(3-bromophenyl)-6-methyl-2-(methylthio)-1,4-
dihydropyrimidine-5-carboxylate (2i)
1
Yellow solid, m.p. 101-104 ºC, 98% yield; H NMR (400 MHz, CDCl ) δ 7.43 (s, 1H),
3
7
.34 (d, J = 7.8 Hz, 1H), 7.25 (d, J = 7.7 Hz, 1H), 7.15 (t, J = 7.8 Hz, 1H), 6.63 (s, 1H),
.67 (s, 1H), 4.17 – 4.05 (m, 2H), 2.41 (s, 3H), 2.31 (s, 3H), 1.21 (t, J = 7.1 Hz, 3H).
5
1
3
C NMR (101 MHz, CDCl ) δ 166.71, 147.29, 130.21, 130.03, 125.78, 122.49, 60.03,
3
-1
1
4.35, 13.62. IR (KBr) (ν, cm ): 3308, 3231, 3092, 3064, 2988, 2927, 1658, 1619,
1
589, 1566, 1489. HRMS (APCI) m/z calculated for C H BrN O S [M+H] 369.0267;
1
5
17
2
2
found 369.0245.
4
.2.2.10.
Ethyl
6-methyl-2-(methylthio)-4-(naphthalen-2-yl)-1,4-
dihydropyrimidine-5-carboxylate (2j)
1
Yellowish solid, m.p. 127-130 ºC, 98% yield; H NMR (400 MHz, CDCl ) δ 7.82 –
3
7
.78 (m, 3H), 7.71 (s, 1H), 7.54 (d, J = 8.5 Hz, 1H), 7.46 – 7.44 (m, 2H), 5.82 (s, 1H),
1
3
4
.09 (q, J = 7.1 Hz, 2H), 2.42 (s, 3H), 2.36 (s, 3H), 1.19 (t, J = 7.1 Hz, 3H). C NMR
(
101 MHz, CDCl ) δ 166.99, 142.16, 133.38, 132.90, 128.32, 128.17, 127.63, 125.97,
3
-1
1
25.73, 125.35, 77.48, 77.16, 76.84, 59.91, 14.32, 13.60. IR (KBr) (ν, cm ): 3326,
3
218, 3051, 2994, 2965, 2953, 2933, 1652, 1474; HRMS (APPI) m/z calculated for
C H N O S 341.1318 [M+H]; found 341.1319.
19
20
2
2
4
.2.2.11.
Ethyl
4-(4-hydroxyphenyl)-6-methyl-2-(methylthio)-1,4-
dihydropyrimidine-5-carboxylate (2k)
1
Beige solid, m.p. 193-195ºC, 90% yield; H NMR (400 MHz, DMSO- d ) δ 9.49 (s,
6
1
H), 9.21 (s, 1H), 7.00 (d, J = 8.5 Hz, 2H), 6.66 (d, J = 7.7 Hz, 2H), 5.40 (s, 1H), 3.99
1
3
(
q, J = 7.0 Hz, 2H), 2.27 (s, 3H), 2.21 (s, 3H), 1.12 (t, J = 7.0 Hz, 3H). C NMR (101

ACCEPTED MANUSCRIPT
-1
MHz, DMSO- d ) δ 166.29, 127.49, 114.82, 58.85, 14.08, 12.47. IR (KBr) (ν, cm ):
6
3
310, 3068, 2992, 2935, 2798, 2725, 2672, 2590, 2513, 2468, 1658, 1609, 1589, 1511.
HRMS (APPI) m/z calculated for C H N O S 307.1111 [M+H]; found 307.1111.
1
5
18
2
3
4
.2.2.12.
7,7-dimethyl-2-(methylthio)-4-(naphthalen-2-yl)-4,6,7,8-
tetrahydroquinazolin-5(1H)-one (7a)
1
Beige solid, m.p. 125-128 ºC, 90% yield. H NMR (200 MHz, DMSO-d ) δ 7.88 – 7.84
6
(m, 3H), 7.65 (s, 1H), 7.53 – 7.41 (m, 3H), 2.44 – 2.35 (m, 5H), 2.22 (d, J = 16.2 Hz,
1
3
1
H), 2.05 (d, J = 15.2 Hz, 1H), 1.02 (s, 3H), 0.92 (s, 3H). C NMR (50 MHz, DMSO-
d6) δ 132.67, 132.13, 127.77, 127.35, 126.03, 125.57, 124.57, 50.44, 32.16, 28.92,
-1
1
2.58. IR (KBr), (ν, cm ): 3249, 3182, 3110, 3076, 2951, 2925, 2888, 2867, 2817,
798, 2731, 1740, 1709, 1656, 1630, 1605, 1487. HRMS (APPI) m/z calculated for
2
C H N OS [M+H] 351.1526; found 351.1525.
21
22
2
4
.2.2.13.
2-(methylthio)-4-(naphthalen-2-yl)-4,6,7,8-tetrahydroquinazolin-5(1H)-
one (7b)
1
Yellowish solid, m.p. 232-234 ºC, 99% yield. H NMR (200 MHz, DMSO-d ) δ 7.90 –
6
7
.83 (m, 3H), 7.65 (s, 1H), 7.52 – 7.45 (m, 3H), 5.64 (s, 1H), 2.51 – 2.45 (m, 2H), 2.38
1
3
(
s, 3H), 2.26 – 2.23 (m, 2H), 1.93 – 1.85 (m, 2H). C NMR (50 MHz, DMSO-d ) δ
6
1
1
1
3
94.99, 142.24, 132.73, 132.20, 128.07, 127.84, 127.36, 126.05, 125.70, 125.38,
-1
24.56, 36.86, 20.89, 12.65. IR (KBr), (ν, cm ): 3245, 3168, 3055, 2933, 2872, 1709,
646, 1626, 1601, 1481. HRMS (APPI) m/z calculated for C H N OS [M+H]
1
9
18
2
23.1213; found 323.1215.