Synthesis, antioxidant and anticholinesterase activities of novel coumarylthiazole derivatives

Article abstract of DOI:10.1016/j.bioorg.2015.02.002

A newly series of coumarylthiazole derivatives containing aryl urea/thiourea groups were synthesized and their inhibitory effects on acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) were evaluated. The result showed that all the synthesized compounds exhibited inhibitory activity to both cholinesterases. Among them, 1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(4-chlorophenyl)thiourea (f8, IC₅₀ = 4.58 μM) was found to be the most active compound against AChE, and 1-(4-fluorophenyl)-3-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e31) exhibited the strongest inhibition against BuChE with IC₅₀ value of 4.93 μM, which was 3.5-fold more potent than that of galantamine. The selectivity of f8 and e31 were 2.64 and 0.04, respectively. In addition, the cupric reducing antioxidant capacities (CUPRAC) and ABTS cation radical scavenging abilities of the synthesized compounds were investigated for antioxidant activity. Among them, f8, f4 and f6 (IC₅₀ = 1.64, 1.82 and 2.69 μM, respectively) showed significantly better ABTS cation radical scavenging ability than standard quercetin (IC₅₀ = 15.49 μM).

Full text of DOI:10.1016/j.bioorg.2015.02.002

Accepted Manuscript
Synthesis, antioxidant and anticholinesterase activities of novel coumarylthia-
zole derivatives
Belma Zengin Kurt, Isil Gazioglu, Fatih Sonmez, Mustafa Kucukislamoglu
PII:
S0045-2068(15)00011-5
DOI:
http://dx.doi.org/10.1016/j.bioorg.2015.02.002
YBIOO 1788
Reference:
Bioorganic Chemistry
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6 November 2014
Please cite this article as: B.Z. Kurt, I. Gazioglu, F. Sonmez, M. Kucukislamoglu, Synthesis, antioxidant and
anticholinesterase activities of novel coumarylthiazole derivatives, Bioorganic Chemistry (2015), doi: http://
dx.doi.org/10.1016/j.bioorg.2015.02.002
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Synthesis, antioxidant and anticholinesterase activities of novel
coumarylthiazole derivatives
Belma Zengin Kurt,a Isil Gazioglu,a Fatih Sonmez*b and Mustafa Kucukislamogluc
aBezmialem Vakif University, Faculty of Pharmacy, Deparment of Analytical and Medicinal Chemistry, 34093,
Istanbul, TURKEY
bSakarya University, Pamukova Vocational High School, 54900, Sakarya, TURKEY
cSakarya University, Faculty of Arts and Science, Department of Chemistry, 54055, Sakarya, TURKEY
*Corresponding author: Present address: Sakarya University, Pamukova Vocational High School, 54900, Sakarya,
TURKEY. Tel.: +90-264-2953378; fax: +90-264-2953679; e-mail address: fsonmez@sakarya.edu.tr (F.Sonmez).
Abstract
A newly series of coumarylthiazole derivatives containing aryl urea/thiourea groups were
synthesized and their inhibitory effects on acetylcholinesterase (AChE) and butyrylcholinesterase
(BuChE) were evaluated. The result showed that all the synthesized compounds exhibited
inhibitory activity to both cholinesterases. Among them, 1-(4-(8-methoxy-2-oxo-2H-chromen-3-
yl)thiazol-2-yl)-3-(4-chlorophenyl)thiourea (f8, IC50= 4.58 µM) was found to be the most active
compound against AChE, and 1-(4-fluorophenyl)-3-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-
2-yl)urea (e31) exhibited the strongest inhibition against BuChE with IC50 value of 4.93 µM,
which was 3.5-fold more potent than that of galantamine. The selectivity of f8 and e31 were 2.64
and 0.04, respectively. In addition, the cupric reducing antioxidant capacities (CUPRAC) and
ABTS cation radical scavenging abilities of the synthesized compounds were investigated for
antioxidant activity. Among them, f8, f4 and f6 (IC50= 1.64, 1.82 and 2.69 µM, respectively)
showed significantly better ABTS cation radical scavenging ability than standard quercetin
(IC50= 15.49 µM).
Keywords: Acetylcholinesterase, antioxidant, butyrylcholinesterase, coumarin, thiazole.
1

1. Introduction
Alzheimer’s disease (AD) is the most common neurodegenerative disease with symptoms of
memory loss, cognition defect and behavioural impairment [1-3]. AD is associated with a
selective loss of cholinergic neurons in the brain and decreasing levels of acetylcholine (ACh)
[4]. The cholinergic system is the earliest and most profoundly affected neurotransmitter system
in AD, with substantial loss of the forebrain, cortex, and hippocampus. This neurotransmitter
together with these brain regions are critical in the acquisition, processing, and storage of
memories and have supported the use of cholinomimetics in the treatment of AD [5]. It is well
known that two forms of cholinesterases coexist ubiquitously throughout the body, i.e.,
acetylcholinesterase (AChE; EC 3.1.1.7) and butyrylcholinesterase (BuChE; EC 3.1.1.8). AChE
is a hydrolase, and its principal biological role is terminating the impulse transmission at
cholinergic synapses by rapid hydrolysis of the neurotransmitter ACh. Butyrylcholinesterase
(BuChE), also known as pseudocholinesterase, is primarily found in plasma, liver, and muscle
tissues. The pharmacological role of BuChE is not yet completely understood. BuChE may have
a compensatory role in the modulation of the hydrolysis of acetylcholine ACh in brain with
degenerative changes. Consequently, BuChE may be target for increasing the cholinergic tone in
AD patients [6,7]. The classical hypothesis of AD is the cholinergic hypothesis, which suggests
that acetylcholinesterase inhibitors (AChEI) could increase the levels of ACh in AD patients
through the inhibition of AChE and, therefore, relieve some symptoms experienced by AD
patients [8]. At the same time, AD is probably associated with multifaceted etiologies and
pathogenic phenomena, and all mechanisms seem to share oxidative stress as a unifying factor,
which is thought to have a causative role in the pathogenesis of AD recently [9]. Based on these
findings, many efforts have been made in the search for potent AChE inhibitors, and a large
number of naturally occurring and synthetic AChE inhibitors such as ambenonium, ensaculine,
huperzine A, physostigmine, and scopoletin have already been reported (Fig. 1) [10-13].
Regarding the X-ray crystallographic structure of AChE from Torpedo californica, three
main binding sites have been determined: (a) the catalytic triad at the bottom of active site
including Ser200, His440 and Glu327; (b) the catalytic anionic site (CAS) at the vicinity of the
catalytic triad consisting of Trp84, Tyr130, Gly199, His441 and His444; (c) peripheral anionic
site (PAS) at the gorge rim comprising Tyr70, Asp72, Tyr121, Trp279 and Tyr334 [14-16].
AChE inhibitors may inhibit AChE via a competitive mechanism, by interacting with the
catalytic active site (CAS) of the enzyme, via a non-competitive mechanism, by binding with the
peripheral anionic site (PAS), or via both mechanisms, by exerting a dual binding AChE
2

inhibition [17].
Several studies have revealed that the tricycle and heterocyclic rings (such as tacrine,
quinolizidinyl and coumarin derivatives) showed strong parallel π-π stacking against CAS, and
the amidic or imidic fragments interact with the catalytic triad of the active site [18-21]. The
coumarin heterocyclic framework is now considered as a privileged structure, which is a common
moiety found in many biologically active natural and therapeutic products and thus represents a
very important pharmacophore [22,23]. Recently, this scaffold has also been reported as an
AChE inhibitor [24]. The various research groups have synthesized amidic- or imidic-based
AChE inhibitors. These studies have reported that a hydrogen bond has been formed between
amid moieties of the inhibitors and anionic sites of the enzyme [25,26]. The urea group was
chosen on the basis of its potency of H-bonding, probability of complexation and wide biological
activity [27-30]. We report here a new hybrid molecule (1) based on frameworks of the coumarin
modified by the addition of thiazole ring (Fig. 2). In the design of this new molecule, the
coumarylthiazole moiety was substituted with urea group contained phenyl ring. We
hypothesized that these heterocyclic and aromatic rings may show strong parallel π-π stacking
against CAS of the enzyme. On the other hand, the presence of urea moiety contributes to
inhibitor activity by interacting with active sites. In this study, a series of 42 novel urea/thiourea
substituted coumarylthiazole derivatives (e1-e34 and f1-f8) were synthesized and their
antioxidant activities and inhibitory effects on AChE and BuChE were evaluated.
2. Results and discussion
2.1. Chemistry
The synthetic procedures employed to obtain the target compounds e1-e34 and f1-f8 are depicted
in Scheme 1. Compounds b1-3 were synthesized from salicylaldehydes by the literature [31],
then they were brominated with molecular bromine in chloroform. 2-amino-4-(R1-coumarin-3-
yl)thiazoles (d1-3) were obtained by the reactions of c1-3 with thiourea. Finally, compounds d1-3
were reacted with arylisocyanates in THF to get product coumarylthiazole containing urea
derivatives (e1-e34), but thiourea derivatives weren’t synthesized by the same procedure.
Although several experiments were carried on different conditions, only 8 new thiourea
substituted coumarylthiazole derivatives (f1-8) were obtained by the reactions of d1-3 with
arylisothiocyanates in DMF at room temperature.
All the new compounds were characterized by 1H NMR, 13C NMR, IR, MS and elemental
analysis. In the infrared spectra of the synthesized compounds, it was possible to observe the
3

absorptions between 3370 and 3250 cm-1 relating to NH stretch for urea derivatives, between
3300 and 3200 cm-1 relating to NH stretch for thiourea derivatives, about 1550 cm-1 relating to
C=N stretch for thiazole, absorptions in about 1680 cm-1 from coumarin carbonyl moiety stretch
and absorptions between 1660 and 1725 cm-1 from urea and thiourea carbonyl moiety stretching.
From the 1H NMR spectra, the resonance due to the hydrogen attached to the amide nitrogen was
between 8.75 and 11.97 ppm. The signals for aromatic hydrogens were observed between 6.90
13
and 8.62 ppm, the signal of proton at thiazole ring was detected 8.55 ppm. From the C NMR
spectra, the signals can be seen 159-161 ppm, relating to coumarin carbonyl and thiazole ring.
This is followed by the sign about 153 ppm for urea carbonyl and about 175 ppm for thiourea
carbonyl.
2.2. Biological activities
2.2.1. Inhibitory activities against AChE and BuChE
The inhibitory activities of the synthesized compounds (e1-e34 and f1-f8) against AChE and
BuChE were performed according to the Ellman’s method using galantamine as the reference
compound [32]. The IC50 values for AChE and BuChE inhibitions are summarized in Table 1.
The results showed that most of the synthesized compounds exhibited inhibitory activity against
ChEs. The IC50 values were between 4.58 and >200 µM for AChE and between 4.93 and 194.55
µM for BuChE inhibitory activity. Among the synthesized compounds, f8 (IC50 = 4.58 µM)
showed the highest inhibitory activity against AChE. Although the potency was less than those of
tacrine (IC50 = 0.086 µM) and galantamine (IC50 = 2.41 µM), it approached to that of
rivastigmine (IC50 = 4.15 µM) [4,20]. e31 exhibited the strongest inhibition against BuChE with
IC50 value of 4.93 µM, which was 3.5-fold more potent than that of galantamine (IC50 = 17.38
µM). Donepezil is currently the most used ChE inhibitors [4]. Although the potency of f8 was
152-fold less than that of donepezil (IC50 = 0.03 µM) against AChE, the potency of e31 was 1.06-
fold less than that of donepezil (IC50 = 4.66 µM) against BuChE.
2.2.2. Structure-activity relationships (SAR)
The following conclusions should be noted regarding the ChEs inhibitory data of Table 1.
(i) The electron-donating group (methoxy) or electron-withdrawing group (nitro) at the
coumarin ring did not cause a regular change on the inhibitory activity.
(ii) The compounds containing thiourea groups have higher the ChE inhibitory activity than
the compounds containing urea groups, while they have same substituents (compared
4

f1 with e8; f2 with e11; f3 with e15; f4 with e16; f5 with e17; f6 with e21; f7 with
e22; f8 with e24).
(iii) Among the thiourea substituted compounds, a halogen atom (F- and Cl-) at the phenyl
ring increased the inhibitory effect for AChE, while bonded the electron-donating
groups (methyl and methoxy) and not bonded any atom to the phenyl ring. This fact
was not observed for BuChE.
(iv) Moving the methoxy group at the phenyl ring from the orto-position to the meta-position
and the para-position led to a major decline of the inhibitory activity for both ChEs
(compared e1 (R2=2'-OCH3, IC50 = 38.02 and 30.32 µM for AChE and BuChE,
respectively) and e2 (R2=3'-OCH3, IC50 = 52.77 and 74.99 µM for AChE and BuChE,
respectively) with e3 (R2=4'-OCH3, IC50 = 95.86 and 82.55 µM for AChE and BuChE,
respectively); compared e14 (R2=3'-OCH3, IC50 = 83.77 and 56.52 µM for AChE and
BuChE, respectively) with e15 (R2=4'-OCH3, IC50 = 119.46 and 99.62 µM for AChE
and BuChE, respectively); compared e27 (R2=3'-OCH3, IC50 = 38.69 and 77.50 µM
for AChE and BuChE, respectively) with e28 (R2=4'-OCH3, IC50 = >200 and 118.50
µM for AChE and BuChE, respectively)).
(v) While R1 was H and 8-OCH3, moving the fluorine atom at the phenyl ring from the orto-
position to the meta-position and the para-position led to a decrease of the inhibitory
activity for AChE (compared e7 (R2=2'-F, IC50 = 31.46 µM for AChE) and e8 (R2=3'-
F, IC50 = 58.05 µM for AChE) with e9 (R2=4'-F, IC50 = 101.13 µM for AChE);
compared e20 (R2=2'-F, IC50 = 66.20 µM for AChE) and e21 (R2=3'-F, IC50 = 67.013
µM for AChE) with e22 (R2=4'-F, IC50 = 115.01 µM for AChE)).
(vi) While R1 was 6-NO2 and 8-OCH3, the halogen series at the para-position of the phenyl
ring showed a qualitative relationship for increasing inhibitory activity with growing
size and polarizability for AChE (for size and polarizability, I > Br > Cl > F, for
inhibitory activity, e26 (R2=4'-I, IC50 = 61.32 µM for AChE) > e25 (R2=4'-Br, IC50 =
69.13 µM for AChE) > e24 (R2=4'-Cl, IC50 = 107.42 µM for AChE) > e22 (R2=4'-F,
IC50 = 115.01 µM for AChE); e34(R2=4'-I, IC50 = 26.73 µM for AChE) > e33 (R2=4'-
Br, IC50 = 46.23 µM for AChE) > e32 (R2=4'-Cl, IC50 = 129.23 µM for AChE) > e31
(R2=4'-F, IC50 = 134.50 µM for AChE)). While R1 was H, this relationship was not
observed for AChE.
Interestingly, the opposite effect was observed against BuChE (e9 (R2=4'-F, IC50 =
84.88 µM for BuChE) > e11 (R2=4'-Cl, IC50 = 88.43 µM for BuChE) > e12 (R2=4'-Br,
5

IC50 = 95.42 µM for BuChE) > e13 (R2=4'-I, IC50 = 111.79 µM for BuChE); e22
(R2=4'-F, IC50 = 63.26 µM for BuChE) > e24 (R2=4'-Cl, IC50 = 70.59 µM for BuChE)
> e25 (R2=4'-Br, IC50 = 116.76 µM for BuChE) > e26 (R2=4'-I, IC50 = 159.29 µM for
BuChE); e31 (R2=4'-F, IC50 = 4.93 µM for BuChE) > e32 (R2=4'-Cl, IC50 = 78.26 µM
for BuChE) > e33 (R2=4'-Br, IC50 = 92.50 µM for BuChE) > e34 (R2=4'-I, IC50 =
97.46 µM for BuChE)).
(vii)
While R1 was H, moving the fluorine atom at the phenyl ring from the orto-position
to the meta-position and the para-position led to a decrease of the inhibitory activity
for BuChE (compared e7 (R2=2'-F, IC50 = 34.47 µM for BuChE) and e8 (R2=3'-F,
IC50 = 62.24 µM for BuChE) with e9 (R2=4'-F, IC50 = 84.88 µM for BuChE)). While
R1 was 8-OCH3, this decrease was not observed for BuChE.
2.2.3. Inhibition mechanism of AChE
The inhibition of AChE can be accomplished irreversibly (organophosphates form a strong
covalent bond with the serine residue in the catalytic triad), pseudoirreversibly (using carbamates,
as the carbamylated serine is slowly hydrolysed to regenerate the active enzyme), or reversibly
(transient noncovalent binding through electrostatic interactions with the active and/or peripheral
sites), depending on the nature of the interaction of the inhibitor with the enzyme site [33]. The
reversible inhibitors may be classified as (a) active-site inhibitors directed toward the catalytic
anionic sub-site at the bottom of the gorge, (b) peripheral anionic site inhibitors which bind at the
entrance to the gorge, or (c) elongated gorge-spanning inhibitors which bridge the two sites [33].
Several crystallographic studies and original computational analysis have demonstrated
that the tricycle ring have interacted strong parallel π-π stacking against the indole ring of Trp84
[14,34-36]. Hydrophobic contacts have also been observed between the ring system and Ser200,
His440 and Phe331. At the mouth of the gorge, the π-π stacking interaction between the phenyl
ring of the compounds and Tyr70 has been found. Additionally, in the middle of the gorge, the π-
π stacking interaction has been observed between the coumarin and the benzene ring of Phe330
[15,16,20,21]. A hydrogen bond has also been formed between amid moieties of the compounds
containing urea, carbamate or sulphonamide groups and Tyr70 and His447 [37-40].
We believe that there are hydrophobic contacts and π-π stacking interactions between the
phenyl, thiazole and coumarin rings of the synthesized compounds and CAS or PAS of AChE.
We also think that a hydrogen bond is formed between urea moieties of them and Tyr70 or
His447.
6

2.3. Antioxidant activity assay
2.3.1. ABTS cation radical scavenging assay
The ABTS method is based on the ability of hydrogen or electron-donating antioxidants to
decolorize the performed radical monocation of 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic
acid) generated due to oxidation of ABTS with potassium persulfate [41]. The results in Table 1
indicated that six synthesized compounds exhibited good radical scavenging ability. Among
them, f8, f4, f6, f7 and f3 (IC50= 1.64, 1.82, 2.69, 3.31 and 5.49 µM, respectively) showed
significantly better activity than quercetin (IC50= 15.49 µM). The compounds (f1-f8) containing
thiourea groups showed significantly better ABTS cation radical scavenging activity than
containing urea groups (e1-e34).
2.3.2. CUPRAC assay
As a distinct advantage over other electron-transfer based assays (e.g., Folin, FRAP, ABTS,
DPPH), CUPRAC is superior in regard to its realistic pH close to the physiological pH,
favourable redox potential, accessibility and stability of reagents, and applicability to lipophilic
antioxidants as well as hydrophilic ones [42]. The cupric reducing antioxidant capacities of the
synthesized compounds (e1-34 and f1-8) were determined according to the literature method [42]
using quercetin as the reference compound. Among the synthesized compounds, only f3 (A0.50
=
16.02 µM) showed better cupric reducing antioxidant activity than quercetin (A0.50 = 18.47 µM).
The others have so much less the cupric reducing antioxidant capacity than quercetin (Table 1).
3. Conclusion
In conclusion, a series of 42 novel urea/thiourea substituted coumarylthiazoles derivatives (e1-
e34 and f1-f8) were synthesized and their antioxidant activities and effects on AChE and BuChE
were evaluated with this study. In addition, structure–activity relationship was presented. All the
synthesized compounds inhibited the AChE and BuChE activities. Among them, f8 was found to
be the most active compound for AChE inhibitory activity, and e31 exhibited the strongest
inhibition against BuChE with IC50 value of 4.93 µM, which was 3.5-fold more potent than that
of galantamine. Additionally, most of the synthesized compounds exhibited good ABTS cation
radical scavenging ability. Among them, only six compounds showed significantly better activity
than quercetin. The SAR study revealed that the inhibitory activity of synthesized compounds
could also be influenced by the type and position of substituent on the phenyl ring.
7

4. Experimental
4.1. General methods
Melting points were taken on a Barnstead Electrothermal 9200. IR spectra were measured on a
1
13
Shimadzu Prestige-21 (200 VCE) spectrometer. H and C NMR spectra were measured on a
1
13
Varian Infinity Plus spectrometer at 300 and at 75 Hz, respectively. H and C chemical shifts
are referenced to the internal deuterated solvent. Mass spectra were obtained using
MICROMASS Quattro LC-MS-MS spectrometer. The elemental analyses were carried out with a
Leco CHNS-932 instrument. Spectrophotometric analyses were performed by a BioTek Power
Wave XS (BioTek, USA). The electric eel acetylcholinesterase (AChE, Type-VI-S, EC 3.1.1.7,
425.84 U/mg, Sigma) and horse serum butyrylcholinesterase (BuChE, EC 3.1.1.8, 11.4 U/mg,
Sigma) were purchased from Sigma (Steinheim, Germany). The other chemicals and solvents
were purchased from Fluka Chemie, Merck, Alfa Easer and Sigma-Aldrich.
4.2. Synthesis procedures and spectral data
3-acetylcoumarin derivatives (b1-3): A mixture of benzaldehyde derivatives (a1-3) (3 mmol),
reactive methylene compound (3 mmol), and L-proline (0.3 mol) was heated at 80 °C for 1 h. The
reaction was monitored by TLC. After completion of the reaction, the mixture was cooled and
recrystallized from ethanol to get pure crystalline b1-3 in 75-80% yields. Spectral data of these
compounds were matched with the literature [31].
3-(bromoacetyl)coumarin derivatives (c1-3): To a solution of 1 mol b1-3 in 20 mL chloroform
was added 1 mol of bromine in 5 mL chloroform, with intermittent shaking and warming to
decompose an addition product. The mixture was heated at 50 °C for 15 min. on a water-bath to
expel most of the hydrogen bromide, then cooled and filtered. The solid was washed with ether
and recrystallized with acetic acid. c1-3 were obtained in 85%, 75%and 80% yields, respectively.
Spectral data of these compounds were matched with the literature [43].
3-(2-amino-1,3-thiazol-4-yl)coumarin derivatives (d1-3): Thiourea (5 mmol) was added to the
solution of c1-3 (5 mmol) in boiling ethanol (20 mL). The mixture was refluxed for 1 hour, then
cooled and neutralized with aqueous ammonia. The precipitate was filtered off, washed with
ethanol and used directly without crystallization or other purification. d1-3 were obtained in 80%,
78% and 75% yields, respectively. Spectral data of these compounds were matched with the
literature [44].
8

Synthesis of coumarylthiazol urea derivatives (e1-34): To a solution of d1-3 (1 mmol) and
triethyl amine (1 mL) in dry THF was added isocyanate derivatives (1 mmol). The mixture was
refluxed for 12 hours with stirring, than cooled and evaporated to dryness. The crude product was
washed with chloroform and dried under vacuum. The products were recrystallized from ethanol
over 98% purity. e1-34 were obtained with 77-99 % yields.
1-(2-methoxyphenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e1)
Yellowish powder, 62% yield, mp. 351-353 °C; IR: 3238, 3126, 1707, 1603, 1529, 1457,1329,
1
1249, 1088, 1026, 753, 693, 577, 457 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.87 (3H, s),
6.89-7.07 (2H, m), 7.35-7.49 (3H, m), 7.59-7.65 (1H, m), 7.85-7.87 (1H, dd, J= 1.46, 7.6 Hz),
7.93 (1H,s), 8.09-8.12 (1H, dd, J= 1.17, 7.9 Hz ), 8.59 (1H, s), 8.83 (1H, s, NH), 11.26 (1H, s,
13
NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 55.6, 108.7, 112.2, 114.5, 114.8, 116.0, 119.8,
120.3, 125.3, 129.5, 131.9, 132.4, 139.0, 140.6, 142.8, 153.5, 155.8, 159.3, 159.6, 160.6; LC-MS
(m/z): 394.06 [MH+]. Anal. Calcd. for C20H15N3O4S: C, 61.06; H, 3.84; N, 10.68; found: C,
61.15; H, 3.80; N, 10.74.
1-(3-methoxyphenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e2)
Yellow powder, 82% yield, mp. 353 °C; IR: 3361, 3299, 1680, 1670, 1544, 1494, 1290, 1197,
1
1178, 1107, 750, 682 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.76 (3H, s), 6.62-6.66 (1H,
dd, , J= 1.8, 8.2 Hz ), 6.99 (1H, d, J= 7.9 Hz), 7.20-7.38 (2H, m), 7.40-7.48 (2H, m), 7.64 (1H, t,
J= 7.3 Hz), 7.88 (1H, d, J= 7.0 Hz), 7.95 (1H, s), 8.61 (1H, s), 9.01 (1H, s, NH), 10.71 (1H, s,
13
NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 55.7, 105.0, 109.0, 111.6, 114.3, 116.5, 119.7,
120.9, 125.4, 129.5, 130.4, 132.5, 139.1, 140.2, 142.6, 151.2 153.0, 159.3, 159.4, 160.4; LC-MS
(m/z): 394.09 [MH+]. Anal. Calcd. for C20H15N3O4S: C, 61.06; H, 3.84; N, 10.68; found: C,
61.09; H, 3.87; N, 10.65.
1-(4-methoxyphenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e3)
Yellowish solid, 80% yield, mp. 353-354 °C; IR: 3350, 3294, 1670, 1665, 1541, 1504, 1290,
1
1232, 1168, 1035, 1109, 748, 665 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.72 (3H, s),
6.90 (2H, d, J= 2.3Hz), 7.34-7.44 (4H, m), 7.58-7.64 (1H, td, J= 1.5, 7.3 Hz), 7.85 (1H, t, J= 6.4
13
Hz), 7.90 (1H, s), 8.57 (1H, s), 8.77 (1H, s, NH), 10.61 (1H, s, NH); C NMR (DMSO-d6, 75
MHz) δ/ppm: 55.8, 114.1, 114.7, 116.5, 119.8, 121.0, 121.3, 125.3, 129.5, 131.9, 132.4, 139.0,
9

142.6, 152.2, 153.0, 155.8, 159.4, 159.5; LC-MS (m/z): 394.06 [MH+]. Anal. Calcd. for
C20H15N3O4S: C, 61.06; H, 3.84; N, 10.68; found: C, 61.12; H, 3.86; N, 10.63.
1-(p-tolyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e4)
Yellow solid, 80% yield, mp. 357-358 °C; IR: 3356, 3287, 1680, 1670, 1541, 1512, 1284, 1242,
1
1178, 1107, 744, 669 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 2.24 (3H, s), 7.11 (2H, d, J=
8.2 Hz), 7.34-7.45 (4H, m), 7.61 (1H, t, J=7.3 Hz), 7.85 (1H, d, J= 6.8 Hz), 7.91 (1H, s), 8.57
(1H, s ), 8.84 (1H, s, NH), 10.63 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 21.0, 114.2,
116.6, 119.4, 119.8, 121.0, 125.4, 129.5, 130.0, 132.4, 132.5, 136.4, 139.0, 142.6, 152.1, 153.0,
159.4, 159.5; LC-MS (m/z): 378.08 [MH+]. Anal. Calcd. for C20H15N3O3S: C, 63.65; H, 4.01; N,
11.13; found: C, 63.68; H, 4.06; N, 11.08.
1-phenyl-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e5)
Yellowish powder, 80% yield, mp. 355-357 °C; IR: 3352, 3288, 1678, 1670, 1544, 1500, 1244, ,
1
1178, 1109, 783, 686 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.06 (1H, t, J= 7.6 Hz), 7.32-
7.45(5H, m), 7.49 (1H, d, J= 8.8 Hz), 7.65 (1H, t, J= 7.2 Hz ), 7.89 (1H, d, J= 6.6 Hz), 7.95 (1H,
s), 8.62 (1H, s), 8.98 (1H, s, NH), 10.71 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm:
114.3, 115.9, 116.6, 119.3, 121.9, 122.7, 123.6, 125.4, 129.6, 132.3, 132.5, 139.0, 142.8, 152.7,
153.0, 159.5, 159.9; LC-MS (m/z): 363.08 [MH+]. Anal. Calcd. for C19H13N3O3S: C, 62.80; H,
3.61; N, 11.56; found: C, 62.85; H, 3.64; N, 11.50.
1-(4-nitrophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e6)
Yellow powder, 76% yield, mp. 356 °C; IR: 3365, 3296, 1664, 1650, 1552, 1498,1289, 1253,
1
1111, 1030, 746, 664 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.36-7.45 (2H, m), 7.62 (1H,
t, J= 8.2 Hz), 7.73 (2H, d, J= 8.5 Hz), 7.85 (1H, d, J= 7.3 Hz), 7.97 (1H, s), 8.20 (2H, d, J= 9.1
13
Hz), 8.59 (1H, s), 9.60 (1H, s, NH), 10.91 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm:
114.2, 116.7, 119.3, 119.7, 121.1, 125.4, 129.4, 130.2, 132.3, 136.3, 139.0, 140.0, 142.3, 152.0,
153.2, 159.3, 159.5; LC-MS (m/z): 409.03 [MH+]. Anal. Calcd. for C19H12N4O5S: C, 55.88; H,
2.96; N, 13.72; found: C, 55.83; H, 2.90; N, 13.76.
1-(2-flourophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e7)
White solid, 80% yield, mp. 358 °C; IR: 3356, 3230, 1685, 1670, 1525, 1504, 1282, 1253, 1203,
1
1089, 750, 694 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.07-7.11 (1H, td, J= 3.5, 5.3 Hz),
10

7.12-7.21 (1H, td, J= 1.2, 7.6 Hz ), 7.25-7.32 (1H, m), 7.36-7.41 (1H, td, J= 1.1, 7.6 Hz ), 7.44
(1H, d, J= 8.5 Hz), 7.59-7.64 (1H, td, J= 1.8, 8.2 Hz), 7.84-7.87 (1H, dd, J= 1.5, 7.9 Hz), 7.95
(1H, s), 8.10-8.16 (1H, td, J= 1.4, 8.2 Hz), 8.60 (1H, s), 9.05 (1H, s, NH), 10.99 (1H, s, NH); 13C
NMR (DMSO-d6, 75 MHz) δ/ppm: 110.0, 114.3, 116.0, 116.6, 119.8, 120.9, 121.5, 125.4, 127.0,
129.6, 131.8, 132.5, 139.1, 142.7, 151.9, 153.1, 158.6, 159.4, 159.7; LC-MS (m/z): 382.09
[MH+]. Anal. Calcd. for C19H12FN3O3S: C, 59.84; H, 3.17; N, 11.02; found: C, 59.92; H, 3.10; N,
11.08.
1-(3-fluorophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e8)
Yellowish powder, 85% yield, mp. 351-353 °C; IR: 3373, 3346, 1716, 1666, 1552, 1487, 1274,
1
1140, 1112, 790, 681 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 6.84-6.89 (1H, td, J= 1.75,
8.2 Hz), 7.16-7.19 (1H, dd, J= 1.17, 78.2 Hz ), 7.31-7.51 (4H, m), 7.59-7.65 (1H, td, J= 1.46, 7.3
Hz), 7.85-7.87 (1H, dd, J= 1.46, 7.61 Hz ), 7.94 (1H, s), 8.59 (1H, s), 9.17 (1H, s, NH), 10.78
13
(1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 111.3, 114.8, 116.2, 116.9, 119.3, 121.1,
121.8, 125.7, 127.2, 129.4, 131.3, 132.1, 139.3, 142.4, 151.5, 152.0, 153.2, 158.3, 159.5; LC-MS
(m/z): 382.08 [MH+]. Anal. Calcd. for C19H12FN3O3S: C, 59.84; H, 3.17; N, 11.02; found: C,
59.80; H, 3.13; N, 11.05.
1-(4-fluorophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e9)
Yellow powder, 86% yield, mp. 355 °C; IR: 3350, 3290, 1668, 1660, 1548, 1487, 1282, 1205,
1
1178, 1103, 750, 657 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.18 (2H, t, J= 6.4 Hz), 7.37-
7.54 (4H, m), 7.61-7.67 (1H, td, J= 1.5, 5.8 Hz), 7.86-7.89 (1H, dd, J= 1.5, 7.9 Hz ), 7.95 (1H, s),
8.60 (1H, s ), 9.00 (1H, s, NH), 10.73 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 114.3,
116.0, 116.3, 116.9, 119.8, 121.3, 121.4, 125.4, 129.5, 130.1, 132.4, 135.4, 139.0, 142.7, 152.2,
153.0, 155.7, 159.4, 160.2; LC-MS (m/z): 382.06 [MH+]. Anal. Calcd. for C19H12FN3O3S: C,
59.84; H, 3.17; N, 11.02; found: C, 59.86; H, 3.12; N, 11.09.
1-(3-chlorophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e10)
Yellow powder, 85% yield, mp. 356-358 °C; IR: 3379, 3294, 1666, 1660, 1546, 1504, 1292,
1
1249, 1178, 1109, 752, 671 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.1 (1H, s), 7.33 (1H,
s), 7.38-7.45 (3H, m), 7.61 (1H, d, J= 6.6 Hz), 7.70 (1H, s), 7.85 (1H, s), 7.94 (1H, s), 8.58 (1H,
13
s), 9.13 (1H, s, NH), 10.79 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 114.3, 115.8,
116.2, 118.4, 119.3, 121.1, 123.5, 125.3, 127.2, 129.4, 130.3, 132.1, 135.3, 139.0, 144.2, 152.0,
11

153.2, 159.3, 159.5; LC-MS (m/z): 398.08 [MH+]. Anal. Calcd. for C19H12ClN3O3S: C, 57.36; H,
3.04; N, 10.56; found: C, 57.30; H, 3.08; N, 10.52.
1-(4-chlorophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e11)
Yellowish powder, 88% yield, mp. 357 °C; IR: 3365, 3288, 1668, 1660, 1544, 1490, 1276, 1240,
1
1178, 1109, 750, 659 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.34-7.43 (5H, m), 7.49
(1H, d, J= 8.5 Hz), 7.60 (1H, t, J= 7.4 Hz), 7.88 (1H, d, J= 7.6 Hz), 7.92 (1H, s), 8.56 (1H, s),
9.06 (1H, s, NH), 10.73 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 114.8, 115.9, 116.4,
118.6, 121.0, 123.7, 127.0, 129.5, 130.2, 132.5, 135.0, 139.2, 144.0, 152.1, 153.0, 159.4, 160.3;
LC-MS (m/z): 398.16 [MH+]. Anal. Calcd. for C19H12ClN3O3S: C, 57.36; H, 3.04; N, 10.56;
found: C, 57.39; H, 3.01; N, 10.58.
1-(4-bromophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e12)
Yellowish powder, 80% yield, mp. 343-345 °C; IR: 3373, 3346, 1666, 1552, 1461, 1274, 1161,
1
1112, 867, 681, 517 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.36-7.51 (6H, m), 7.60-7.65
(1H, td, J= 1.75, 8.4 Hz), 7.87 (1H,d, J= 7.9 Hz), 7.95 (1H, s), 8.59 (1H, s), 9.10 (1H, s, NH),
13
10.77 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 105.0, 111.6, 112.4, 118.6, 120.7,
121.1, 125.3, 131.4, 139.3, 140.8, 142.9, 146.1, 152.0, 156.7, 158.1, 159.5, 160.3; LC-MS
(m/z): 441.96 [MH+]. Anal. Calcd. for C19H12BrN3O3S: C, 51.60; H, 2.73; N, 9.50; found: C,
51.65; H, 2.76; N, 9.46.
1-(4-iodophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e13)
Yellowish powder, 78% yield, mp. 326-328 °C; IR: 3361, 3281, 1673, 1605, 1539, 1483, 1376,
1
1180, 812, 747, 584 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.22-7.49 (4H, m), 7.56-7.65
(3H,m), 7.85 (1H,d, , J= 7.8 Hz ), 7.93 (1H, s), 8.58 (1H, s), 9.09 (1H, s, NH), 10.74 (1H, s, NH);
13C NMR (DMSO-d6, 75 MHz) δ/ppm: 103.8, 111.6, 114.3, 114.8, 120.1, 121.7, 125.3, 132.5,
139.2, 140.7, 142.9, 145.8, 152.0, 156.8, 158.5, 159.5, 160.9; LC-MS (m/z): 489.97 [MH+]. Anal.
Calcd. for C19H12IN3O3S C, 46.64; H, 2.47; N, 8.59 found: C, 46.60; H, 2.41; N, 8.55.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(3-methoxyphenyl)urea (e14)
Yellow powder, 92% yield, mp. 320 °C; IR: 3352, 3300, 1681, 1670, 1543, 1496, 1288, 1219,
1
1172, 1105, 773, 694 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.73 (3H, s), 3.90 (3H, s),
6.61 (1H, d, J= 7.4 Hz), 6.96 (1H, d, J= 7.9 Hz), 7.16-7.37 (5H,m), 7.92 (1H, s), 8.53 (1H, s),
12

13
8.95 (1H, s, NH), 10.66 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 55.7, 56.7, 105.0,
111.6, 114.4, 114.6, 120.3, 120.7, 121.1, 125.3, 130.4, 139.2, 140.2, 142.3, 142.6, 146.9, 152.0,
156.5, 159.1, 159.3, 160.4; LC-MS (m/z): 424.13 [MH+]. Anal. Calcd. for C21H17N3O5S C,
59.57; H, 4.05; N, 9.92; found: C, 59.60; H, 4.10; N, 9.87.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(4-methoxyphenyl)urea (e15)
Yellowish powder, 90% yield, mp. 338 °C; IR: 3348, 3290, 1678, 1660, 1541, 1504, 1278, 1222,
1
1178, 1109, 777, 640 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.72 (3H, s), 3.92 (3H, s),
6.90 (2H, d, J= 7.8 Hz), 7.27-7.33 (2H, m), 7.37 (3H, d, J= 8.8 Hz ), 7.90 (1H, s), 8.53 (1H, s),
13
8.74 (1H, s, NH), 10.57 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 55.8, 56.8, 114.3,
116.2, 119.8, 121.5, 122.4, 125.4, 129.4, 132.8, 135.1, 139.0, 142.3, 146.4, 152.3, 153.1, 156.8,
159.4, 159.5; LC-MS (m/z): 424.12 [MH+]. Anal. Calcd. for C21H17N3O5S C, 59.57; H, 4.05; N,
9.92; found: C, 59.54; H, 4.08; N, 9.95.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(p-tolyl)urea (e16)
Yellowish powder, 86% yield, mp. 337 °C; IR: 3388, 3282, 1680, 1670, 1539, 1508, 1273, 1240,
1
1176, 1093, 773, 673 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 4.24 (3H, s), 3.91 (3H, s),
7.12 (2H, d, J= 8.5 Hz), 7.27-7.29 (2H, m), 7.32-7.37 (3H, m), 7.90 (1H, s), 8.52 (1H, s), 8.82
(1H, s, NH), 10.60 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 24.2, 56.7, 111.6, 114.3,
114.7, 120.4, 121.7, 125.4, 129.5, 132.0, 139.0, 140.3, 142.3, 146.8, 152.0, 156.7, 158.3, 159.4,
159.5; LC-MS (m/z): 408.14 [MH+]. Anal. Calcd. for C21H17N3O4S C, 61.90; H, 4.21; N, 10.31;
found: C, 61.96; H, 4.25; N, 10.25.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-phenylurea (e17)
Yellowish powder, 82% yield, mp. 338 °C; IR: 3365, 3304, 1681, 1670, 1541, 1492, 1290, 1246,
1
1182, 1103, 758, 696 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.92 (3H, s), 7.06 (1H, d, J=
6.7 Hz), 7.29-7.37 (6H, m), 7.47 (2H, d, J= 7.6 Hz), 8.55 (1H, s), 8.94 (1H, s, NH), 10.66 (1H, s,
13
NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 56.7, 111.7, 114.2, 116.8, 119.7, 121.5, 122.3,
125.4, 126.4, 129.7, 132.3, 139.0, 140.1, 142.3, 152.0, 156.8, 159.4, 159.7; LC-MS (m/z): 394.18
[MH+]. Anal. Calcd. for C20H15N3O4S C, 61.06; H, 3.84; N, 10.68 ; found: C, 61.00; H, 3.89; N,
10.65.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(3-nitrophenyl)urea (e18)
13

Yellow powder, 76% yield, mp. 339 °C; IR: 3350, 3296, 1668, 1660, 1529, 1508, 1342, 1249,
1
1176, 1109, 777, 659 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.92 (3H, s), 7.29-7.39 (3H,
m) 7.60 (1H, t, J= 8.2 Hz ), 7.78 (1H, d, J= 7.9 Hz), 7.88 (1H, d, J= 7.3 Hz), 7.96 (1H, s), 8.53
(2H, d, J=6.2 Hz), 9.42 (1H, s, NH), 10.81 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm:
56.8, 111.8, 113.0, 114.3, 114.8, 116.7, 119.8, 120.5, 121.8, 125.4, 129.7, 132.2, 139.0, 140.2,
141.2, 142.0, 152.3, 156.7, 159.4, 159.5; LC-MS (m/z): 439.29 [MH+]. Anal. Calcd. for
C20H14N4O6S C, 54.79; H, 3.22; N, 12.78; found: C, 54.74; H, 3.28; N, 12.75.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(4-nitrophenyl)urea (e19)
Yellow powder, 74% yield, mp. 345 °C; IR: 3361, 3302, 1676, 1670, 1552, 1496, 1342, 1253,
1
1178, 1107, 777, 688 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.92 (3H, s), 7.31 (2H, d, J=
4.9 Hz), 7.49 (1H, s), 7.73 (2H, d, J= 9.1 Hz ), 7.98 (1H, s), 8.21 (2H, d, J= 8.8 Hz), 8.55 (1H, s),
9.60 (1H, s, NH), 10.91 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 56.7, 111.7, 114.2,
116.4, 119.8, 121.4, 125.5, 129.8, 132.2, 139.0, 140.3, 141.5, 142.4, 144.0, 152.3, 156.7, 159.3,
159.5; LC-MS (m/z): 439.18 [MH+]. Anal. Calcd. for C20H14N4O6S C, 54.79; H, 3.22; N, 12.78;
found: C, 54.83; H, 3.20; N, 12.76.
1-(2-fluorophenyl)-3-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e20)
White solid, 78% yield, mp. 337-338 °C; IR: 3350, 3234, 1701, 1680, 1544, 1508, 1280, 1253,
1118, 1089, 773, 673 cm-1; 1H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.91 (3H, s), 7.05-7.15 (1H,
m), 7.20 (1H, t, J= 8.0 Hz), 7.29-7.39 (4H, m), 7.94 (1H, s), 8.14 (1H, t, J= 8.2 Hz ), 8.55 (1H, s),
9.04 (1H, s, NH), 10.96 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 56.7, 111.7, 114.3,
116.5, 116.7, 117.5, 119.6, 122.4, 123.7, 125.7, 126.2, 132.3, 139.0, 141.3, 142.0, 152.0, 156.7,
158.1, 159.4, 159.5; LC-MS (m/z): 412.04 [MH+]. Anal. Calcd. for C20H14FN3O4S C, 58.39; H,
3.43; N, 10.21; found: C, 58.42; H, 3.45; N, 10.18.
1-(3-fluorophenyl)-3-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e21)
Yellowish powder, 78% yield, mp. 335-338 °C; IR: 3358, 3290, 1673, 1612, 1463, 1257, 1107,
1
970, 771, 677 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.94 (3H, s), 6.85-6.91 (1H, td, J=
2.34, 8.49 Hz), 7.20 (1H, d, J= 7.3 Hz), 7.32-7.43 (4H, m), 7.49 (1H, d, J= 9.6 Hz ), 7.97 (1H, s),
8.58 (1H, s), 9.18 (1H, s, NH), 10.80 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 56.5,
111.5, 113.2, 116.8, 117.7, 118.3, 119.2, 121.4, 123.9, 125.5, 127.2, 132.5, 139.2, 141.3, 142.5,
14

152.1, 156.8, 158.6, 159.3, 159.6; LC-MS (m/z): 412.08 [MH+]. Anal. Calcd. for C20H14FN3O4S
C, 58.39; H, 3.43; N, 10.21; found: C, 58.35; H, 3.40; N, 10.25.
1-(4-fluorophenyl)-3-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e22)
Yellowish powder, 90% yield, mp. 342 °C; IR: 3356, 3296, 1672, 1660, 1548, 1506, 1274, 1257,
1
1193, 1099, 775, 651 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.94 (3H, s), 7.15-7.22 (2H,
m), 7.31-7.51 (3H, m), 7.52-7.55 (2H, m), 7.95 (1H, d, J= 4.1 Hz), 8.57 (1H, d, J= 4.4 Hz ), 9.01
(1H, s, NH), 10.74 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 56.8, 111.8, 114.2, 116.5,
117.8, 119.8, 122.3, 125.7, 126.3, 130.0, 131.8, 136.1, 138.2, 140.1, 142.9, 146.5, 152.1, 157.8,
159.5, 159.7; LC-MS (m/z): 412.23 [MH+]. Anal. Calcd. for C20H14FN3O4S C, 58.39; H, 3.43; N,
10.21; found: C, 58.33; H, 3.40; N, 10.27.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(3-chlorophenyl)urea (e23)
Yellowish powder, 84% yield, mp. 337-338 °C; IR: 3390, 3260, 1676, 1669, 1548, 1508, 1271,
1
1242, 1190, 1103, 777, 673 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.94 (3H, s), 7.10-7.13
(1H, dd, J= 2.0, 6.1 Hz), 7.31-7.39 (4H, m), 7.40 (1H, d, J= 2.9 Hz), 7.72 (1H, d, J= 1.5 Hz), 7.96
13
(1H, d, J= 2.4 Hz), 8.57 (1H, s), 9.14 (1H, s, NH), 10.83 (1H, s, NH); C NMR (DMSO-d6, 75
MHz) δ/ppm: 56.5, 111.7, 114.2, 116.5, 116.8, 119.8, 121.2, 122.3, 125.7, , 128.7, 130.0, 132.1,
135.3, 139.0, 140.1, 142.2, 152.0, 156.7, 159.7, 159.9; LC-MS (m/z): 428.13 [MH+]. Anal.
Calcd. for C20H14ClN3O4S C, 56.14; H, 3.30; N, 9.82; found: C, 56.18; H, 3.35; N, 9.78.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(4-chlorophenyl)urea (e24)
Yellowish powder, 94% yield, mp. 346 °C; IR: 3379, 3286, 1670, 1665, 1546, 1510, 1274, 1242,
1
1176, 1101, 775, 636 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.93 (3H, s), 7.33 (3H,s),
7.38 (2H, d, J= 8.2 Hz), 7.53 (2H, d, J= 8.2 Hz), 7.96 (1H, s), 8.57 (1H, s), 9.16 (1H, s, NH),
13
10.85 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 56.8, 111.8, 114.3, 116.7, 119.8,
121.0, 122.3, 125.5, 127.2, 128.7, 132.1, 135.0, 139.0, 142.4, 152.1, 156.5, 159.3, 159.5; LC-MS
(m/z): 428.11 [MH+]. Anal. Calcd. for C20H14ClN3O4S C, 56.14; H, 3.30; N, 9.82;found: C,
56.12; H, 3.33; N, 9.85.
1-(4-bromophenyl)-3-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e25)
Cream powder, 76% yield, mp. 294-296 °C; IR: 3363, 3277, 1673, 1607, 1543, 1466, 1272,
1
1238, 1102, 819, 774 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.90 (3H, s), 7.28-7.50 (7H,
15

13
m), 7.93 (1H, s), 8.53 (1H, s), 9.07 (1H, s, NH), 10.74 (1H, s, NH); C NMR (DMSO-d6, 75
MHz) δ/ppm: 56.7, 111.3, 113.2, 115.9, 116.7, 118.8, 121.6, 123.3, 128.5, 132.6, 135.5, 138.7,
140.5, 142.8, 152.3, 156.5, 159.4, 159.8; LC-MS (m/z): 472.01 [MH+]. Anal. Calcd. for
C20H14BrN3O4S C, 50.86; H, 2.99; N, 8.90; found: C, 50.82; H, 2.95; N, 8.95.
1-(4-iodophenyl)-3-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e26)
Yellow powder, 62% yield, mp. 281-283 °C; IR: 3359, 3305, 1678, 1602, 1539, 1478, 1272,
1
1234, 1103, 778, 627 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.94 (3H, s), 7.28-7.43 (4H,
m), 7.58-7.67 (3H, m), 7.96 (1H, s), 8.57 (1H, s), 9.07 (1H, s, NH), 10.76 (1H, s, NH); 13C NMR
(DMSO-d6, 75 MHz) δ/ppm: 56.5, 110.8, 113.2, 116.1, 117.3, 119.2, 122.0, 124.7, 128.3, 131.0,
132.5, 139.6, 140.7, 142.9, 152.0, 156.7, 159.5, 159.9; LC-MS (m/z): 519.95 [MH+]. Anal.
Calcd. for C20H14IN3O4S C, 46.26; H, 2.72; N, 8.09; found: C, 46.28; H, 2.74; N, 8.05.
1-(3-methoxyphenyl)-3-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e27)
Dark yellow powder, 60% yield, mp. 340-342 °C; IR: 3361, 3291, 1727, 1695, 1606, 1551, 1490,
1
1338, 1208, 1082, 826, 782 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.74 (3H, s), 6.60-6.63
(1H, dd, J= 2.05, 8.2 Hz), 6.95-6.97 (1H, dd, J= 1.46, 7.61 Hz), 7.15-7.24 (2H, m), 7.66 (1H, d,
J= 9.07 Hz ), 7.98 (1H, s), 8.37-8.40 (1H, dd, J=2.92, 9.07 Hz), 8.72 (1H, s), 8.89 (1H, d, J= 2.92
13
Hz ), 9.02 (1H, s, NH), 10.61 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 55.7, 104.9,
109.0, 111.5, 115.6, 118.1, 118.6, 120.3, 122.5, 125.3, 126.7, 130.5, 137.7, 140.2, 142.1, 144.4,
152.0, 156.5, 159.4, 160.4; LC-MS (m/z): 439.09 [MH+]. Anal. Calcd. for C20H14N4O6S C,
54.79; H, 3.22; N, 12.78; found: C, 54.75; H, 3.26; N, 12.76.
1-(4-methoxyphenyl)-3-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e28)
Dark yellow powder, 60% yield, mp. 328-330 °C; IR: 3364, 3140, 1733, 1698, 1612, 1553, 1343,
1
1242, 1089, 834, 744 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.72 (3H, s), 6.90 (2H, d, J=
9.07 Hz), 7.38 (2H, d, J= 8.78 Hz), 7.66 (1H, d, J= 9.37 Hz), 7.98 (1H, s), 8.38-8.41 (1H,dd,
J=2.63, 9.37 Hz), 8.72 (1H, s), 8.83 (1H, s, NH), 8.89 (1H, d, J= 2.92 Hz), 10.58 (1H, s, NH); 13C
NMR (DMSO-d6, 75 MHz) δ/ppm: 55.8, 103.8, 109.2, 111.3, 115.8, 118.9, 121.3, 122.8, 125.7,
132.5, 137.3, 140.2, 142.0, 144.8, 152.2, 156.7, 159.5, 160.3; LC-MS (m/z): 439.10 [MH+].
Anal. Calcd. for C20H14N4O6S C, 54.79; H, 3.22; N, 12.78; found: C, 54.73; H, 3.24; N, 12.75.
1-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(p-tolyl)urea (e29)
16

Dark yellow powder, 60% yield, mp. 347-348 °C; IR: 3355, 3132, 1690, 1606, 1530, 1345, 1237,
1
1177, 1102, 1003, 82, 638 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 2.24 (3H, s), 7.10 (2H,
d, J= 8.2 Hz), 7.33 (2H, d, J= 8.2 Hz), 7.61 (1H, d, J= 9.07 Hz), 7.93 (1H, s), 8.35 (1H, d, J=
9.07 Hz), 8.65 (1H, s, NH), 8.85 (2H, d, J= 12.59 Hz), 10.54 (1H, s, NH); 13C NMR (DMSO-d6,
75 MHz) δ/ppm: 24.5, 104.2, 108.2, 111.0, 115.5, 118.3, 120.3, 122.4, 125.3, 132.0, 138.6, 140.5,
142.8, 149.0, 152.3, 156.8, 159.7, 160.4; LC-MS (m/z): 423.10 [MH+]. Anal. Calcd. for
C20H14N4O5S C, 56.87; H, 3.34; N, 13.26; found: C, 56.85; H, 3.30; N, 13.30.
1-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-phenylurea (e30)
Dark yellow powder, 64% yield, mp. 351-352 °C; IR: 3360, 3295, 1727, 1694, 1604, 1542, 1482,
1
1339, 1290, 995, 743 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.06 (1H, t, J= 7.32 Hz),
7.34 (2H, t, J= 7.32 Hz), 7.49 (2H, d, J= 7.32 Hz), 7.67 (1H, d, J= 9.07 Hz ), 8.01 (1H,s), 8.39-
8.42 (1H, dd, J= 2.63, 9.07 Hz ), 8.74 (1H, s), 8.91 (1H, d, J= 2.92 Hz), 9.03 (1H, s, NH), 10.65
13
(1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 104.3, 108.2, 111.7, 114.3, 118.5, 122.3,
123.7, 132.6, 135.3, 139.2, 142.1, 144.3, 145.2, 152.1, 156.3, 158.5, 159.7; LC-MS (m/z):
409.07 [MH+]. Anal. Calcd. for C19H12N4O5S C, 55.88; H, 2.96; N, 13.72; found: C, 55.85; H,
2.93; N, 13.76.
1-(4-fluorophenyl)-3-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e31)
Dark yellow powder, 56% yield, mp. 349-350 °C; IR: 3365, 3298, 1734, 1680, 1615, 1551, 1342,
1
1294, 1205, 1089, 839, 640 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.18 (2H, t, J= 8.78
Hz), 7.48-7.57 (2H, m), 7.67 (1H, d, J= 9.37 Hz), 8.00 (1H, s), 8.22 (1H, d, J= 9.37 Hz ), 8.73
13
(1H, s), 8.90 (1H, s), 9.04 (1H, s, NH), 10.66 (1H, s, NH); C NMR (DMSO-d6, 75 MHz)
δ/ppm: 114.5, 116.3, 118.5, 120.6, 122.3, 125.6, 126.8, 128.1, 129.0, 136.5, 138.4, 142.0, 144.2,
152.0, 156.6, 158.7, 159.5; LC-MS (m/z): 427.13 [MH+]. Anal. Calcd. for C19H11FN4O5S C,
53.52; H, 2.60; N, 13.14; found: C, 53.55; H, 2.63; N, 13.10.
1-(4-chlorophenyl)-3-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e32)
Yellow powder, 62% yield, mp. 348-350 °C; IR: 3358, 3285, 1739, 1693, 1540, 1486, 1339,
1
1284, 1085, 834, 610 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.38 (2H, d, J= 9.66 Hz),
7.52 (2H, d, J= 9.07 Hz), 7.66 (1H, d, J= 9.07 Hz), 8.00 (1H, s), 8.38-8.41 (1H, dd, J= 2.63, 9.07
13
Hz ), 8.72 (1H, s), 8.89 (1H, d, J= 2.63 Hz), 9.16 (1H, s, NH), 10.71 (1H, s, NH); C NMR
(DMSO-d6, 75 MHz) δ/ppm: 115.7, 118.1, 120.3, 120.8, 122.5, 125.3, 126.7, 127.1, , 129.5,
17

137.7, 138.0, 142.1, 144.4, 152.0, 156.5, 158.5, 159.3; LC-MS (m/z): 443.06 [MH+]. Anal.
Calcd. for C19H11ClN4O5S C, 51.53; H, 2.50; N, 12.65; found: C, 51.57; H, 2.53; N, 12.60.
1-(4-bromophenyl)-3-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e33)
Yellowish powder, 84% yield, mp. 334-337 °C; IR: 3345, 3250, 1744, 1607, 1522, 1351, 1259,
1
1093, 829, 506 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.41-7.51 (4H, m), 7.65 (1H, d, J=
9.07 Hz), 8.00 (1H, s), 8.37-8.40 (1H, dd, J= 2.63, 9.07 Hz), 8.72 (1H, s), 8.87 (1H, s), 9.14 (1H,
13
s, NH), 10.70 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 115.3, 1185, 121.2, 121.8,
122.8, 126.4, 127.8, , 129.2, 132.5, 137.3, 138.8, 142.7, 144.5, 152.1, 156.7, 158.3, 159.6; LC-
MS (m/z): 486.98 [MH+]. Anal. Calcd. for C19H11BrN4O5S C, 46.83; H, 2.28; N, 11.50; found:
C, 46.87; H, 2.32; N, 11.46.
1-(4-iodophenyl)-3-(4-(6-nitro-2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea (e34)
Yellow powder, 75 % yield, mp. 331-333 °C; IR: 3354, 3246, 1741, 1698, 1605, 1519, 1479,
1
1349, 1265, 828 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.33 (2H, d, J= 8.78 Hz), 7.63-
7.68 (3H, m), 8.00 (1H, s), 8.38-8.41 (1H, dd, J= 2.63, 9.07 Hz), 8.72 (1H, s), 8.89 (1H, d, J=
13
2.63 Hz ), 9.12 (1H, s, NH), 10.70 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 113.7,
116.1, 120.7, 121.8, 123.9, 127.5, 129.8, 131.5, 132.3, 137.3, 138.4, 142.5, 144.9, 152.5, 155.8,
158.6, 159.8; LC-MS (m/z): 534.97 [MH+]. Anal. Calcd. for C19H11IN4O5S C, 42.71; H, 2.08; N,
10.49; found: C, 42.76; H, 2.04; N, 10.46.
Synthesis of coumarylthiazol thiourea derivatives (f1-8): To a solution of d1-3 (1 mmol) and
triethyl amine (1 mL) in DMF was added isothiocyanate derivatives (1 mmol). The mixture was
stirred at temperature for 12 hours and then poured into cold 1 M HCl. The precipitate was
filtered and washed with cold water. The crude products were recrystallized from ethanol over
99% purity. f1-8 were obtained with 77-99 % yields.
1-(3-fluorophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)thiourea (f1)
Yellow powder, 74 %yield, mp. 348 °C; IR: 3373, 3346, 1678, 1666, 1552, 1487, 1274, 1199,
1
1097, 756, 678cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 6.89 (1H, t, J= 8.2 Hz), 7.20 (1H, d,
J= 7.9 Hz), 7.33-7.53 (4H, m), 7.65 (1H, t, J= 7.9 Hz), 7.89 (1H, d, J= 7.6 Hz), 7.97 (1H, s), 8.62
13
(1H, s), 9.25 (1H, s, NH), 10.84 (1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 110.0,
114.4, 115.1, 116.5, 119.8, 120.9, 122.8, 125.3, 126.3, 129.5, 131.1, 132.5, 139.0, 142.7, 152.1,
18

159.1, 159.4, 166.7, 175.9; LC-MS (m/z): 398.04 [MH+]. Anal. Calcd. for C19H12FN3O2S2 C,
57.42; H, 3.04; N, 10.57; found: C, 57.46; H, 3.08; N, 10.51.
1-(4-chlorophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)thiourea (f2)
Yellow powder, 70% yield, mp. 287 °C; IR: 3321, 3289, 1705, 1680, 1544, 1489, 1238, 1180,
1
1089, 754, 651 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 7.38-7.54 (4H, m), 7.63-7.71 (3H,
m), 7.83 (1H, d, J= 7.9 Hz), 7.95 (1H, s), 8.60 (1H, s), 10.77 (1H, s, NH), 11.95 (1H, s, NH); 13C
NMR (DMSO-d6, 75 MHz) δ/ppm: 114.2, 119.8, 120.1, 122.5, 122.8, 125.4, 126.2, 127.3, 129.2,
131.2, 132.5, 139.0, 142.3, 152.1, 159.2, 159.4, 174.8; LC-MS (m/z): 414.07 [MH+]. Anal. Calcd.
for C19H12ClN3O2S2 C, 55.13; H, 2.92; N, 10.15; found: C, 55.18; H, 2.90; N, 10.11.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(4-methoxyphenyl)thiourea (f3)
Orange powder, 74% yield, mp. 249-251 °C; IR: 3350, 3272, 1724, 1680, 1542, 1506, 1273,
1
1188, 1089, 775, 651cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.96 (3H, s), 3.99 (3H, s),
6.92-7.00 (2H, m), 7.14 (1H, s), 7.21-7.49 (2H, m), 7.54 (2H, d, J= 6.7 Hz), 7.87 (1H, s), 8.48
(1H, s), 10.52 (1H, s, NH), 11.69 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 55.9, 56.7,
114.2, 114.5, 114.6, 120.8, 121.2, 125.2, 125.3, 126.3, 131.8, 139.2, 142.3, 146.9, 157.7, 159.0,
159.1, 161.0, 174.3; LC-MS (m/z): 440.20 [MH+]. Anal. Calcd. for C21H17N3O4S2 C, 57.39; H,
3.90; N, 9.56; found: C, 57.35; H, 3.94; N, 9.58.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(p-tolyl)thiourea (f4)
Cream powder, 72% yield, mp. 353 °C; IR: 3324, 3289, 1720, 1680, 1573, 1506, 1251, 1190,
1
1091, 779, 698cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 2.30 (3H, s), 3.92 (3H, s), 7.20 (2H,
d, J= 8.2 Hz), 7.31 (3H, t, J= 7.3 Hz), 7.50 (2H, d, J= 8.2 Hz), 7.90 (1H, s), 8.52 (1H, s), 10.65
(1H, s, NH), 11.78 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 24.2, 56.7, 114.2, 114.6,
120.1, 122.3, 125.3, 126.2, 128.7, 129.0, 131.3, 134.3, 139.2, 142.0, 146.5, 157.8, 159.5, 160.1,
175.5; LC-MS (m/z): 424.11 [MH+]. Anal. Calcd. for C21H17N3O3S2 C, 59.56; H, 4.05; N, 9.92;
found: C, 59.51; H, 4.09; N, 9.90.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-phenylthiourea (f5)
Yellowish powder, 72% yield, mp. 355 °C; IR: 3350, 3289, 1724, 1680, 1575, 1510, 1255, 1192,
1
1089, 773, 684 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.91 (3H, s), 7.21-7.49 (6H, m),
13
7.65 (2H, d, J= 7.3 Hz), 7.91 (1H, s), 8.53 (1H, s), 10.68 (1H, s, NH), 11.85 (1H, s, NH); C
19

NMR (DMSO-d6, 75 MHz) δ/ppm: 56.5, 114.1, 114.5, 120.2, 122.4, 124.2, 125.3, 126.7, 128.3,
129.0, 134.5, 139.0, 142.2, 146.4, 157.2, 159.4, 160.0, 175.8; LC-MS (m/z): 410.02 [MH+]. Anal.
Calcd. for C20H15N3O3S2 C, 58.66; H, 3.69; N, 10.26; found: C, 58.63; H, 3.65; N, 10.30.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(3-fluorophenyl)thiourea (f6)
Cream powder, 74% yield, mp. 354 °C; IR: 3322, 3204, 1726, 1702, 1577, 1469, 1282, 1195,
1091, 773, 682 cm-1; 1H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.94 (3H, s), 7.06 (1H, s), 7.32-7.42
(5H, m), 7.76 (1H, d, J= 10.08 Hz), 7.96 (1H, s), 8.57 (1H, s), 10.80 (1H, s, NH), 11.97 (1H, s,
13
NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 56.7, 114.1, 114.5, 116.2, 116.8, 120.2, 125.7,
126.8, 128.2, 129.0, 130.2, 134.6, 139.0, 142.2, 146.0, 157.3, 159.2, 160.1, 161.2, 176.9; LC-MS
(m/z): 428.11 [MH+]. Anal. Calcd. for C20H14FN3O3S2 C, 56.19; H, 3.30; N, 9.83; found: C,
56.24; H, 3.35; N, 9.78.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(4-fluorophenyl)thiourea (f7)
Cream powder, 72% yield, mp. 333-336 °C; IR: 3330, 3203, 1724, 1700, 1554, 1504, 1280,
1
1197, 1091, 781, 680 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.97 (3H, s), 7.11-7.40 (5H,
m), 7.42-7.51 (1H, m), 7.61-7.66 (1H, m), 7.90 (1H, d, J= 3.5 Hz), 8.53 (1H, d, J= 3.5 Hz ), 9.74
(1H, s, NH), 11.07 (1H, s, NH); 13C NMR (DMSO-d6, 75 MHz) δ/ppm: 56.5, 114.1, 114.5, 116.2,
120.2, 125.5, 126.7, 128.3, 129.0, 134.6, 139.0, 142.1, 146.3, 157.5, 158.0, 159.5, 160.1, 176.8;
LC-MS (m/z): 428.06 [MH+]. Anal. Calcd. for C20H14FN3O3S2 C, 56.19; H, 3.30; N, 9.83; found:
C, 56.20; H, 3.32; N, 9.80.
1-(4-(8-methoxy-2-oxo-2H-chromen-3-yl)thiazol-2-yl)-3-(4-chlorophenyl)thiourea (f8)
Yellow powder, 73% yield, mp. 268 °C; IR: 3350, 3292, 1714, 1700, 1489,1475, 1249, 1190,
1
1089, 779, 700 cm-1; H NMR (DMSO-d6, 300 MHz) δ/ppm: 3.91 (3H, s), 7.30 (3H, s), 7.45
(2H, d, J= 6.2 Hz), 7.69 (2H, d, J= 6.1 Hz), 7.90 (1H, s), 8.51 (1H, s), 10.72 (1H, s, NH), 11.92
13
(1H, s, NH); C NMR (DMSO-d6, 75 MHz) δ/ppm: 56.7, 114.0, 114.2, 120.1, 122.5, 125.3,
126.5, 128.2, 129.0, 130.4, 134.6, 139.0, 142.0, 146.1, 157.5, 159.3, 159.9, 176.5; LC-MS (m/z):
445.07 [MH+]. Anal. Calcd. for C20H14ClN3O3S2 C, 54.11; H, 3.18; N, 9.47; found: C, 54.17; H,
3.22; N, 9.43.
20

4.2. Anticholinesterase activity assays
Acetyl- (AChE) and butyryl-cholinesterase (BuChE) inhibitory activities of the synthesized
compounds were determined according to the literature method [32]. In this study, the IC50 of a
substance have been determined by constructing an absorbance and/or inhibition (%) curve and
examining the effect of different concentrations. IC50 values have been calculated for a given
antagonist by determining the concentration needed to inhibit half of the maximum biological
response of the agonist. The substrates of the reaction were acetylthiocholine iodide and
butyrylthiocholine iodide. 5,5'-dithio-bis(2-nitrobenzoic) acid (DTNB) was used for the
measurement of the anticholinesterase activity. The solutions of the samples and galantamine in
n-propanol were prepared at concentration of 4000 µg/mL. Aliquots of 150 µL of 100 mM
phosphate buffer (pH 8.0), 10 μL of sample solution and 20 μL AChE (2.476x10-4 U/μL) (or
3.1813x10-4 U/μL BChE) solution were mixed and incubated for 15 min at 25ºC. 10 μL of DTNB
solution was prepared by adding 2.0 mL of pH 7.0 and 4.0 mL of pH 8.0 phosphate buffers to a
mixture of 1.0 mL of 16 mg/mL DTNB and 7.5 mg/mL NaHCO3 in pH 7.0 phosphate buffers.
The reaction was initiated by the addition of 10 μL (7.1 mM) acetylthiocholine iodide (or 0.79
mM butyrylthiocholine iodide). In this method, the activity was measured by following the
yellow colour produced as a result of the thio anion produced by reacting the enzymatic
hydrolysis of the substrate with DTNB. Propyl alcohol was used as a solvent to controls. The
hydrolysis of these substrates was monitored using a BioTek Power Wave XS at 412 nm. The
results were calculated as IC50.
4.3. Antioxidant activity assays
In CUPRAC assay, the absorbance values were used to calculate for A0.50, but in ABTS assay,
inhibition (%) values were used to calculate for IC50.
ABTS cation radical decolorization assay: ABTS·+ scavenging activities of the synthesized
compounds were determined according to the literature method [41]. The solution of ABTS·+
radical was generated by dissolving 19.2 mg of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic
acid) (7 mM ABTS) and 3.3 mg K2S2O3 in distilled water (5 mL). The solution was kept in dark
for 24 hours at room temperature, and the absorbance of the solution was fixed to ~0.70 at 734
nm by dilution. The solutions of the samples were prepared in n-propanol at a concentration of
1000 µg/mL. The absorbance was measured in room temperature at 734 nm, after 6 minutes from
ABTS·+ addition. The decrease in the absorption was used to calculate. The results were
calculated as IC50. Propyl alcohol was used as a solvent to controls.
21

Cupric reducing antioxidant capacity assay (CUPRAC): Cupric reducing antioxidant capacities
of the synthesized compounds were determined according to the literature method [42]. The
solutions of compounds and standards were prepared in n-propanol at a concentration of 1000
µg/mL. Different volumes (1000 mg/L and 54.5 mL) of the sample were added to a solution
prepared by adding 61.0 µL of 10 mM CuCl2, 61.0 µL 7.5 mM neocuproine and 61.0 µL of 1.0
mM NH4CH3COO buffer (pH 7), respectively. The absorbance was measured in room
temperature at 450 nm, after an hour. The results were calculated as A0.50. Propyl alcohol was
used as a solvent to controls.
Acknowledgments
This work was supported by the Sakarya Research Fund of the Sakarya University. Project
Number: 2014-50-02-003.
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24

Figures, Tables and schemes captions
Figure 1. Structures of well-known cholinesterase inhibitors.
Figure 2. Design strategy of the targeted compounds.
Table 1. In vitro inhibition IC50 and A0.50 values (µM) and selectivity of compounds e1-e34 and
f1-f8 for AChE and BuChE and antioxidant activities.
Scheme 1. Synthesis of new urea/thiourea substituted coumarylthiazole derivatives.
25

Schemes:
Scheme 1. Synthesis of new urea/thiourea substituted coumarylthiazole derivatives.
26

Tables:
Table 1. In vitro inhibition IC50 and A0.50 values (µM) and selectivity of compounds e1-e34 and f1-f8 for
AChE and BuChE and antioxidant activities.
8
8
5
2
O
O
2
O
O
7
7
R1
6
R1
3
3
N
N
13
6
13
11
15
NH
11
15
NH
17
5
4
4
17
NH
1'
NH
1'
S
2'
S
2'
O
S
3'
3'
6'
e1-e34
6'
f1-f8
4'
4'
5'
R2
5'
R2
AChE
BuChE
Selectivity
indexb
0.80
1.42
0.86
0.67
1.02
0.70
1.10
1.07
0.84
0.60
2.32
0.79
1.23
0.67
0.83
1.16
0.92
0.87
1.80
1.20
1.64
0.55
0.60
0.66
1.69
2.60
2.00
<0.59
2.07
0.82
0.04
0.60
2.00
3.65
1.27
3.18
0.76
0.70
0.47
0.98
0.50
2.64
ABTS·+
CUPRAC
A0.50 (µM)c
>200
>200
>200
>200
>200
>200
147.37± 0.01
>200
Compound
R1
R2
IC50 (µM)a
IC50 (µM)a
IC50 (µM)a
96.09 ± 3.38
107.97± 1.82
166,89 ± 0,67
97.42± 1.22
182.50± 2.92
61.50 ± 2.92
>200
H
H
H
H
H
H
H
H
H
H
H
H
2'-OCH3
3'-OCH3
4'-OCH3
4'-CH3
H
4'-NO2
2'-F
3'-F
38.02 ± 0.71
52.77 ± 0.77
95.86 ± 3.03
81.45 ± 1.18
73.19 ± 1.38
74.78 ± 3.04
31.46 ± 0.74
58.05 ± 3.24
30.32 ± 1.55
74.99 ± 3.24
82.55 ± 2.01
54.76 ± 0.98
74.65 ± 0.54
52.69 ± 0.99
34.47 ± 2.14
62.24 ± 3.42
84.88 ± 2.67
66.62 ± 0.78
88.43 ± 5.12
95.42 ± 0.13
e1
e2
e3
e4
e5
e6
e7
e8
85.90 ± 1.61
74,92 ± 2,21
170,17 ± 5,74
87,62 ± 1,52
97,49 ± 2,42
>200
>200
>200
94,17 ± 0,2
>200
>200
4'-F
101.13 ± 0.13
111.20 ± 0.81
38.14 ± 3.85
121.22 ± 1.81
91.17 ± 0.05
83.77 ± 0.16
119.46 ± 0.23
82.10 ± 0.98
84.09 ± 0.6
90.04 ± 2.78
46.50 ± 0.49
66.20 ± 1.19
67.013 ± 2.8
115.01 ± 0.74
104.85 ± 1.77
107.42 ± 1.09
69.13 ± 5.05
61.32 ± 0.76
38.69 ± 0.85
>200
93.94 ± 3.01
102.77 ± 0.3
134.50 ± 0.89
129.23 ± 0.13
46.23 ± 1.21
26.73 ± 2.25
16.42 ± 7.11
7.63 ± 2.77
41.18 ± 5.12
51.86 ± 5.96
46.05 ± 0.16
18.40 ± 3.54
23.28 ± 2.7
4.58 ± 0.83
-
192.2 ± 3.56
>200
e9
3'-Cl
4'-Cl
4'-Br
4'-I
3'-OCH3
4'-OCH3
4'-CH3
H
3'-NO2
4'-NO2
2'-F
3'-F
4'-F
3'-Cl
4'-Cl
4'-Br
4'-I
3'-OCH3
4'-OCH3
4'-CH3
H
e10
e11
e12
e13
e14
e15
e16
e17
e18
e19
e20
e21
e22
e23
e24
e25
e26
e27
e28
e29
e30
e31
e32
e33
e34
f1
108.52 ± 2.21
113.46 ± 3.76
127.41 ± 2.63
112.18± 0.01
199.45± 0.06
105.33 ± 1.89
>200
>200
>200
>200
>200
H
111.79 ± 5.52
56.52 ± 0.29
99.62 ± 2.23
95.59 ± 0.57
77.5 ± 4.85
78.44 ± 4.32
83.69 ± 2.6
79.61 ± 3.1
109.72 ± 0.42
63.26 ± 1.05
62.93 ± 0.59
70.59 ± 0.25
116.76 ± 4.04
159.29 ± 2.90
77.5 ± 0.96
118.5 ± 0.52
194.55 ± 0.49
84.51 ± 0.31
4.93 ± 0.56
78.26 ± 0.67
92.5 ± 1.11
97.46 ± 0.66
20.84 ± 0.88
24.30 ± 1.02
31.18 ± 3.03
36.49 ± 1.14
21.81 ± 0.91
17.99 ± 0.41
46.12 ± 0.54
12.09 ± 7.50
-
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
6-NO2
6-NO2
6-NO2
6-NO2
6-NO2
6-NO2
6-NO2
6-NO2
H
>200
177,88 ± 0,56
95,64 ± 0,75
177,79 ± 4,66
78,78 ± 2,42
84,89 ± 2,86
85,306 ± 0,78
60.49 ± 4.22
54,66 ± 0,63
57,58 ± 1,13
74,41 ± 1,58
79,57 ± 2,98
42.72 ± 2.27
65,6 ± 3,02
90.65 ± 2.17
77,57 ± 0,02
27.34 ± 5.08
22.71 ± 0.42
5.49 ± 0.74
1.82 ± 0.70
7.42 ± 1.13
2.69 ± 1.35
3.31± 1.92
1.64 ± 0.78
15.49 ± 2.33
-
>200
>200
>200
160.87 ± 1.85
102.00 ± 1.62
119.49 ± 0.03
131.88 ± 0.02
76.44 ± 0.01
86.09 ± 0.03
92.15 ± 0.01
69.81 ± 0.01
129.51 ± 0.01
104.82 ± 0.01
63.69 ± 0.01
55.87 ± 0.02
16.02 ± 0.15
41.65 ± 0.01
41.06 ± 0.01
55.95 ± 0.02
39.97 ± 0.04
25.73 ± 0.05
18.47± 0.04
-
4'-F
4'-Cl
4'-Br
4'-I
3'-F
H
4'-Cl
4'-OCH3
4'-CH3
H
3'-F
4'-F
4'-Cl
-
-
f2
f3
f4
f5
f6
f7
f8
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
8-OCH3
-
Quercetin
Galantamine
-
2.41 ± 0.12
17.38 ± 0.56
7.21
Donepezild
-
-
0.03±0.0005
4.66±0.503
-
-
Tacrined
-
-
-
-
0.086±0.0049
4.15±0.21
0.013±0.001
0.037±0.01
-
-
-
-
Rivastigminee
a IC50 values represent the means ± S.E.M. of three parallel measurements (p< 0.05).
b Selectivity index = IC50 (BuChE) / IC50 (AChE).
c A0.50 values represent the means ± S.E.M. of three parallel measurements (p< 0.05).
d From ref. [4]
e From ref. [20]
27

Figures:
Fig. 1. Structures of well-known cholinesterase inhibitors.
Fig. 2. Design strategy of the targeted compounds.
28

Graphical abstract;
29