Aryl hydrazones linked thiazolyl coumarin hybrids as potential urease inhibitors

Article abstract of DOI:10.1007/s13738-021-02377-8

Aryl hydrazones bearing thiazolyl coumarin hybrids 1–32 were prepared by following 'one-pot' two-steps reaction scheme. Various arylaldehydes were reacted to thiosemicarbazide under acidic condition to form aryl thiosemicarbazone intermediates which in turn treated with 3-bromoacetyl coumarin under basic condition to afford thiazolyl coumarin hybrids 1–32. All hybrids were recognized by EI- and HREI-MS and ¹H- and ¹³C-NMR spectroscopic techniques. Compounds 1–32 were screened for in vitro inhibitory activity against urease enzyme and displayed good to moderate inhibitory potential in the ranges of IC₅₀ = 16.29 ± 1.1–256.30 ± 1.4?μM. Worth stating that compound 21 (IC₅₀ = 16.29 ± 1.1?μM) was identified as more potent urease inhibitor than the standard acetohydroxamic acid (IC₅₀ = 27.0 ± 0.5?μM). Derivatives 19 (IC₅₀ = 77.67 ± 1.5?μM) and 30 (IC₅₀ = 71.21 ± 1.6?μM) were found to be moderately active. Structure–activity relationship revealed that -F, -Cl, -OH, and -OMe groups and their respective positions on aryl ring are playing important role in urease enzyme inhibition. Molecular docking studies identified important interaction between the ligand (active hybrids) and urease active site.

Full text of DOI:10.1007/s13738-021-02377-8

Journal of the Iranian Chemical Society
https://doi.org/10.1007/s13738-021-02377-8
ORIGINAL PAPER
Aryl hydrazones linked thiazolyl coumarin hybrids as potential urease
inhibitors
Uzma Salar1,3 · Bakhtawer Qureshi1 · Khalid Mohammed Khan1,2,4 · Muhammad Arif Lodhi5 · Zaheer Ul‑Haq3 ·
Farman Ali Khan5 · Fouzia Naz1 · Muhammad Taha4 · Shahnaz Perveen6 · Shafqat Hussain7
Received: 8 March 2021 / Accepted: 7 August 2021
© Iranian Chemical Society 2021
Abstract
Aryl hydrazones bearing thiazolyl coumarin hybrids 1–32 were prepared by following 'one-pot' two-steps reaction scheme.
Various arylaldehydes were reacted to thiosemicarbazide under acidic condition to form aryl thiosemicarbazone intermedi-
ates which in turn treated with 3-bromoacetyl coumarin under basic condition to aford thiazolyl coumarin hybrids 1–32.
All hybrids were recognized by EI- and HREI-MS and 1H- and 13C-NMR spectroscopic techniques. Compounds 1–32 were
screened for in vitro inhibitory activity against urease enzyme and displayed good to moderate inhibitory potential in the
ranges of IC50 =16.29 1.1–256.30 1.4 µM. Worth stating that compound 21 (IC50 =16.29 1.1 µM) was identifed as more
potent urease inhibitor than the standard acetohydroxamic acid (IC50 =27.0 0.5 µM). Derivatives 19 (IC50 =77.67 1.5 µM)
and 30 (IC50 =71.21 1.6 µM) were found to be moderately active. Structure–activity relationship revealed that -F, -Cl,
-OH, and -OMe groups and their respective positions on aryl ring are playing important role in urease enzyme inhibition.
Molecular docking studies identifed important interaction between the ligand (active hybrids) and urease active site.
Keywords Hydrazones · Thiazolyl coumarin · Urease inhibition · In vitro · In silico · Structure–activity relationship
Introduction
* Uzma Salar
uzma_salar_sheikh@hotmail.com
Metalloenzyme urease (EC 3.5.1.5) belongs to amidohydro-
* Khalid Mohammed Khan
lases and possesses two nickel atoms in its active site. It is
broadly distributed in bacteria, algae, fungi, plants, animals,
and other organisms. Being the frst crystallized and puri-
fed from plant Canavalia ensiformis (jack bean) in 1926 by
James B. Sumner, it has noticeable position in enzymology.
It plays indispensable role to catalyze the hydrolysis of urea
into carbon dioxide and ammonia [1–6]. Urea is quite stable
molecule and its urease catalyzed hydrolysis is 1014 times
quicker than un-catalyzed reaction [7]. Hyperactivity of ure-
ase increases the ammonia formation thus increases the pH
in stomach. The increase in alkalinity gives the favorable
condition for the growth and colonization of Helicobac-
ter pylori, a pathogenic bacteria which ultimately leads to
pathologies including peptic, gastric, duodenal ulcers, and
cancer. Hyperactivity of urease is also a virulence factor
in the pathological conditions such as pyelonephritis, uro-
lithiasis, urinary catheter encrustation, hepatic coma, and
hepatic encephalopathy [8–12]. In agriculture sector, urea
is usually utilized as nitrogen fertilizer. On hydrolysis, it
produces ammonia which then further converted into the
khalid.khan@iccs.edu
Farman Ali Khan
drkhalidhej@gmail.com
1
H. E. J. Research Institute of Chemistry, International Center
for Chemical and Biological Sciences, University of Karachi,
Karachi 75270, Pakistan
2
Pakistan Academy of Sciences, 3-Constitution Avenue G-5/2,
Islamabad, Pakistan
3
Dr. Panjwani Center for Molecular Medicine and Drug
Research, International Center for Chemical and Biological
Sciences, University of Karachi, Karachi 75270, Pakistan
4
Department of Clinical Pharmacy, Institute for Research
and Medical Consultations (IRMC), Imam Abdulrahman Bin
Faisal University, P.O. Box 31441, Dammam, Saudi Arabia
5
Department of Biochemistry, Abdul Wali Khan University,
Mardan, KP, Pakistan
6
PCSIR Laboratories Complex, Shahrah-e-Dr. Salimuzzaman
Siddiqui, Karachi 75280, Pakistan
7
Department of Chemistry, University of Baltistan, Skardu,
Gilgit-Baltistan, Pakistan
Vol.:(0123456789)
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Journal of the Iranian Chemical Society
ammonium ions. Plants used these ammonium ions under
ideal soil conditions (pH 6–6.5). Urease hyperactivity during
urea fertilization leads to the liberation of excessive ammo-
nia into the atmosphere and thus deprives plants from vital
nutrients. Furthermore, it surges the ammonia toxicity in
atmosphere and ultimately brings out economic and envi-
ronmental destruction [13–15]. Consequently, it is still desir-
able to identify new, stable, potent, and non-cytotoxic urease
inhibitors which could be an efective strategy to solve prob-
lems triggered by urease enzyme.
antiviral, antituberculosis, and pesticidal activities [32–34].
Thiazole scafold is also well reported for urease inhibition
[35, 36]. Our research team has explored diversely substi-
tuted coumarin thiazole hybrids for their inhibitory potential
against α-glucosidase and α-amylase. These inhibitors were
also found to be potent ROS (reactive oxygen species, NO
(nitric oxide), DPPH, and ABTS inhibitors [37–39]. We have
reported bis-coumarin analogs as potential urease inhibi-
tors [40, 41]. Bis-coumarin bearing OH and X (halogens)
were identifed as potential leads for urease inhibition [41].
Urease inhibitory potential of thiazole ring and the pharma-
cophoric details of bis-coumarin encouraged us to explore
aryl hydrazones bearing thiazolyl coumarin hybrids 1–32
for their possible urease inhibitory activity. Worth noting
that the synthetic library 1–32 is decorated with a range of
diferent substituents including OH and X (halogens). Thus,
this piece of research work demonstrated the synthesis, spec-
troscopic characterization, urease inhibitory screening, and
in silico studies (Fig. 1).
Coumarin is always the center of interest for medici-
nal chemist due to its indispensable and diverse spectrum
of pharmacological potential and less toxicity. Coumarin
is among the most studied scafolds due to its benefcial
impact on human health [16–18]. It is an important class
of compounds possesses diverse range of biological poten-
tials such as anticancer, antioxidant, anti-HIV, anticoagu-
lant, anti-infammatory, antibacterial, anti-leishmanial, and
dyslipidemic activities [19–28]. Apart from diverse bioac-
tivities, cytotoxicity of coumarins was extensively studied
[17, 29]. A number of marketed drugs have coumarin as
a privileged scafold [14]. Warfarin is the best example of
coumarin-based drug, globally used to prolong the survival
of cancer patients [30]. Over the decades, coumarin-based
compounds, natural and synthetic, have shown their poten-
tial toward various bioactivities. Scientist have adopted sev-
eral chemical strategies to link diverse heterocycles with
coumarin scafold in order to identify potential drug candi-
dates for various ailments [17, 31].
Results and discussion
Chemistry
'One-pot' two-steps reaction strategy was adopted to synthe-
size aryl hydrazones linked thiazolyl coumarin hybrids 1–32.
Commercially available arylaldehydes were frst treated
with thiosemicarbazide to aford intermediate (aryl thio-
semicarbazone derivatives). This condensation reaction was
catalyzed by glacial acetic acid and carried out in ethanol
under refux condition for 2 h. Intermediate formation was
confrmed by TLC (thin layer chromatography) monitoring.
Heterocyclic thiazole ring is a privileged scafold of a
range of medicinally important molecules. It is associated
with interesting bioactivities including anticancer, antitumor,
anti-infammatory, analgesic, anticonvulsant, antimicrobial,
O
O
O
O
O
O
S
S
R4
S
R4
N
N
N
N
R2
N
H
N
R2
N
H
HN
R1 R3
R1 R3
N
R2
O
R1
-Amylase Inhibitors
α
-Glucosidase Inhibitors [37]
DPPH and ABTS Radical Scavengers [38]
ROS and NO Inhibitors [39]
α
(A)
(B)
(C)
O
O
O
O
O
O
S
N
R
N
R1
R1
N
H
H
OH
OH
1-32
R2
Non Cytotoxic Urease Inhibitors [41]
IC50 = 16.29 1.1 - 256.30 1.4 µM
Urease Inhibitory Activity
Fig. 1 Bioactive coumarin thiazole hybrids (A, B, C) and aryl hydrazones bearing thiazolyl coumarin hybrids 1–32 as potential urease inhibitors
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Journal of the Iranian Chemical Society
On complete disappearance of starting materials, reaction
was supplemented with 3-bromoacetyl coumarin and fur-
ther refux for 3 h to obtain aryl hydrazones linked thiazolyl
coumarin hybrids 1–32 and TEA (triethylamine) was used to
catalyze the reaction. TEA was used in slight excess to neu-
tralize acetic acid (Scheme 1). Structures of all hybrids 1–32
were identifed by EI-, HREI-MS, 1H-, and 13C-NMR spec-
troscopic techniques. Till to date, compounds 3–6, 14–23,
25, 27–29, 32 are structurally new while other derivatives
are known [42–48].
(IC50 = 27.0 0.5 µM). Its activity may compare with
another analog 16 (not active) having same substitution
pattern but -Cl instead of -F, a complete loss of inhibitory
activity observe. It gives a perception that more electron-
egative -F is worth important than less electronegative
-Cl for urease inhibitory activity. Another aspect is the
size diference of both atoms, as -Cl is big in size than
-F which might cause difculty to bind efectively with
the active site of urease enzyme. Activity comparison of
compound 21 (IC50 = 16.29 1.1 µM) with compound 20
(not active) having -Br instead of -OH showed complete
loss of activity which indicates the active participation of
-OH might be by making hydrogen bonding with enzyme’s
active site. In addition to that, -Br being big in size might
be not allowing the molecule to interact with the active
site. Interestingly, comparing the inhibitory potential of
compound 20 (not active) with 19 (IC50 = 77.67 1.5 µM)
demonstrates the importance of substituent’s position,
such as switching positions of -Br and -F, which brought
moderate inhibitory activity. In this case, compound might
attain the conformation which interacts with the active
pocket of enzyme. Furthermore, compounds 14–17 having
diferent patterns of -Cl and -OH revealed diferent inhibi-
tory attributes. Such as compound 16 was found to be com-
pletely inactive; only the repositioning of -Cl and -OH in
compound 15 (IC50 = 215.23 1.5 µM) showed moderate
inhibitory strength. Unfortunately, switching the positions
of both groups as in compound 14 (not active) brought a
complete loss of activity. It gives an insight that meta and
para positions for -OH and -Cl, respectively, are crucial
for demonstrating the inhibitory strength. It also confrmed
by compound 17, having -OH at ortho and two -Cl groups
at meta positions, and was found inactive. Mono-chloro-
substituted compounds 11 (IC50 = 151.74 2.1 µM) and
12 (IC50 = 221.19 1.6 µM) having -Cl groups at para
and meta, respectively, shown that -Cl at para position
is more actively playing its role as compared to meta
position. Activity of analog 11 can be compared with 13
(IC50 = 177.34 1.9 µM), where addition of another -Cl
group showed decline in the inhibitory strength (Fig. 2).
Enzyme inhibitory activities In vitro
Synthesized aryl hydrazones linked thiazolyl coumarin
hybrids 1–32 were screened for their possible urease inhibi-
tory activity in vitro. A number of analogs 1–3, 5–9, 11–13,
15, 18, 19, 21, 23, 24, 29–32 revealed signifcantly good to
moderate inhibitory potential as shown by IC50 ranges from
16.29 1.1 to 256.30 1.4 µM. Remarkably, compound 16
(IC50 =16.29 1.1 µM) was found exclusively more potent
than standard (acetohydroxamic acid; IC50 =27.0 0.5 µM).
Compounds 19 (IC50 = 77.67 1.5 µM) and 30
(IC50 =71.21 1.6 µM) showed moderate inhibitory poten-
tial (Table 1).
It is pertinent to state that several structural features of
these molecules including coumarin, thiazole, imine group,
NH, and varyingly substituted aryl ring must taking part in
the inhibition process. However, structure–activity relation-
ship (SAR) can only be discussed by focusing on the varying
features such as substitutions on aryl ring. Thus, SAR is
rationalized by analyzing the efect of diferent groups on
inhibitory potential.
Structure–activity relationship (SAR)
Halogen bearing compounds 11–17, 19–21 showed
‘potent’ to ‘not active’ inhibitory potential. Worth reveal-
ing that compound 21 (IC50 = 16.29 1.1 µM) bearing
-F and -OH, para to each other was identifed as most
potent analog of this series on comparison with standard
Scheme 1 Syntheses of aryl
hydrazones linked thiazolyl cou-
marin hybrids 1–32
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Journal of the Iranian Chemical Society
Table 1 In vitro urease inhibitory activities of synthetic aryl hydrazones linked thiazolyl coumarin hybrids 1–32
Urease inhibitory activity
Comp.
1
R
IC50 SEMa (µM)
160.21 0.9
166.34 2.1
76.41 0.1
2
3
NAb
4
190.89 1.8
179.11 1.2
5
6
256.30 1.4
161.45 1.3
187.12 0.1
NAb
7
8
9
10
151.74 2.1
11
12
221.19 1.6
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Journal of the Iranian Chemical Society
Table 1 (continued)
177.34 1.9
13
14
15
NAb
215.23 1.5
NAb
NAb
16
17
18
19
20
21
178.10 1.6
77.67 1.5
NAb
16.29 1.1
NAb
22
23
177.25 1.1
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Journal of the Iranian Chemical Society
Table 1 (continued)
181.52 0.6
24
NAb
NAb
NAb
25
26
27
NAb
28
29
131.61 1.4
71.21 1.6
30
31
141.81 0.6
134.20 0.4
27.0 0.5
32
Acetohydroxamic Acidc
SEMa (Standard error mean); NAb (not active); acetohydroxamic acidc (standard inhibitor of urease enzyme)
Compounds 1–10 bearing -OMe and combinations of
-OMe with -Cl, -OEt, -OAc, and -OH showed comparable
to moderate urease inhibitory strength. Such as deriva-
tive 3 (IC50 = 76.41 0.1 µM) having -OMe and -Cl at
adjacent positions exhibited almost threefold less inhibi-
tory strength on comparison with acetohydroxamic acid
(IC50 = 27.0 0.5 µM). Its activity may be compared with
compound 2 (IC50 =166.34 2.1 µM) in the absence of -Cl
and displayed more than twofold decline in the inhibitory
strength. This comparison shows that -Cl being electron-
egative atom involved in inhibitory potential of molecule.
Interestingly, compound 1 (IC50 = 160.21 0.9 µM) with
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Journal of the Iranian Chemical Society
Fig. 2 Activity comparisons of analogs 11–17 and 19–21
no substitutions on phenyl ring showed activity almost
similar to compound 2. Apparently, it gives a perception
that there is no worth of -OMe substitution but it might
be possible that compound 1 occupied a conformation
which ftted well in the active site. Activity of analog 3 as
compared to analog 5 (IC50 = 190.89 1.8 µM) revealed
that displacement of -Cl group with -OEt brought more
than twofold decreased inhibitory strength. This compari-
son further shows the importance of -Cl group especially
at ortho position. Comparing activities of compounds 5
(IC50 = 190.89 1.8 µM) and 6 (IC50 = 179.11 1.2 µM)
revealed that switching the positions of -OEt and -OMe
in compound 6 brought a slightly increased inhibitory
strength. Compound 8 (IC50 = 161.45 1.3 µM) having
m-OMe and p-OH (adjacent to each other) showed mod-
erate urease inhibitory power. Replacing p-OH with p-
OMe in compound 9 (IC50 = 187.12 0.1 µM) presented
slight decline in activity, whereas replacing p-OH with
p-OAc in compound 4 (not active) showed complete loss
of inhibitory strength. It shows that in this case masked
-OH in the form of -OMe and -OAc is not as good as -OH
for inhibitory activity. Furthermore, -OH group is capa-
ble of making interesting hydrogen bonding interaction
with nickel ions in active pocket of enzyme. Compound
10 bearing 3-OMe groups was found to be completely in
active (Fig. 3).
Compounds 30–32 bearing naphthol, indole, and
thiophene rings showed considerable urease inhibitory
strength. Among them, 2-naphthol bearing derivative 30
(IC50 = 71.21 1.6 µM) showed more than twofold less
inhibitory power when compared to acetohydroxamic acid
(IC50 = 27.0 0.5 µM). Replacing 2-naphthol ring with
indole and thiophene rings as in cases of compounds 31
(IC50 = 141.81 0.6 µM) and 32 (IC50 = 134.20 0.4 µM)
displayed twofold decreased inhibition. Good activity
by the compound 30 might be due to -OH group along
with extended conjugation which facilitates the bind-
ing with the active site of urease enzyme or –OH might
be solely making hydrogen bonding with the active site of
enzyme. Compounds 29 (IC50 = 131.61 1.4 µM) and 23
(IC50 = 177.25 1.1 µM) bearing -OBn along with -OMe
displayed moderate inhibitory strength while solely -OBn
substituted analogs 27 and 28 were found to be completely
inactive. Activity comparison of compounds 23 with 28 and
29 with 27 revealed that -OMe group might playing role in
exhibiting the urease inhibitory potential (Fig. 4).
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Journal of the Iranian Chemical Society
O
O
O
O
H
H
NH
N
HN
N
EtO
O
O
O
O
4
5
OMe
OAc
MeO
Not Active
IC50 = 190.89 1.8 µM
H
H
NH
N
H
HN
N
MeO
EtO
2
OMe
6
IC50 = 166.34 2.1 µM
IC50 = 179.11 1.2 µM
O
O
O
O
O
O
H
NH
N
NH
N
H
HN
N
Cl
3
1
OMe
9
MeO
MeO
IC50 = 76.41 0.1 µM
IC50 = 160.21 0.9 µM
IC50 = 187.12 0.1 µM
O
O
O
O
O
O
H
NH
N
H
HN
N
H
NH
N
OH
OMe
OMe
10
Not Active
MeO
HO
8
IC50 = 161.45 1.3 µM
7
MeO
IC50 = 256.30 1.4 µM
OMe
Fig. 3 Activity comparisons of analogs 1–10
Fig. 4 Activity comparisons of analogs 23, 27–32
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Journal of the Iranian Chemical Society
As a whole, it may have deduced from SAR that groups
like -F, -Cl, -OH, and -OMe playing important role in exhi-
bition of the inhibitory potential; however, nature, size, and
positions of these groups are also playing a crucial role.
Molecular docking studies
Molecular docking were performed to observe the urease
inhibition by understanding the plausible ligand binding
mechanism of synthetic compounds (Fig. 5).
The top-ranked conformer of coumarin-based hybrids
and standard (acetohydroxamic acid) was selected based on
docking score. The docking scores of the compounds 21, 3,
7, and acetohydroxamic acid were -12.38, -10.06, -8.517,
and -6.15 kcal/mol, respectively. Urease binding site is situ-
ated inside the cavity of its internal space where HAE mol-
ecule chelates with nickel ions (Ni798 and Ni799) by means
of hydroxyl oxygen. Important amino acid residues in the
catalytic site are His137, His139, Ala170, Lys220, His249,
His275, Gly280, Cys322, His323, His324, Arg339, Ala363,
Asp363, His137, His139, and KCX220.
Fig. 6 The predicted binding interactions of compounds 21 in the
active site
interaction was observed for compound 21 with Lys169,
Glu166, His324, His322, and KCX220.
The docking study showed that the acetohydroxamic acid
and all the ligands ft into the urease binding pocket. Interac-
tions of the most potent derivative 21 (IC50 =16.29 1.1 μM)
displayed in Fig. 6. Structure of the docking conformation
demonstrated that the potent activity of derivative 21 is due
to making a stable complex of two nickel atoms with nitro-
gen of hydrazinyl group at a distance of 2.99 and 3.20 Å.
The compound was further stabilized by mediating hydrogen
bond interaction via oxygen of coumarin ring and hydroxy-
benzylidene with Met367 and Arg339 at a distance of 1.841
and 3.30 Å, respectively. In addition, the van der Waals
The interaction pattern of analogs 3 and 7 having mod-
erate and least activity, respectively, revealed that ligation
with Ni is vital for stabilization of ligand-enzyme complex.
Figure 7a shows that derivative 3 (IC50 =76 1.1 µM) with
a methoxy group ligated strongly with Ni798 at a bind-
ing distance of 2.23 Å, whereas Ni799 moved away from
methoxy oxygen at the distance of 6.37 Å and this might
be the reason for moderate activity. Similarly, thiazole oxy-
gen is pointed toward Cys322 formed hydrogen bond at a
distance of 1.869 Å. Considering the ftness value, deriva-
tive 3 showed a binding energy -10.06 kcal/mol. Figure 7b
Fig. 5 The location HAE in the
active site of urease, close-up
representation of the active site,
the HAE co-crystallized, and
the corresponding re-docked
form are represented in pink and
cyan color, respectively
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Journal of the Iranian Chemical Society
Fig. 7 The predicted binding interactions of compounds 3 and 7 in the active site of Bacillus pasteurii urease
clearly shows that the position of derivative 7 is far away
from nickel bi-center at the distance of 5.127 and 5.324 Å
and showed no metal ligation, though it formed hydrogen
bond with Lys169. The binding energy of -8.517 kcal/mol
could be the probable clarifcation for the poor in vitro activ-
ity of derivative 7.
(TLC) was carried out on pre-coated silica gel GF-254 alu-
minum plates (Kieselgel 60, 20 20, 0.5 mm thick, E. Merck,
Germany). Spots on TLC plates were visualized under
ultraviolet at 254 nm and 366 nm or iodine vapors. Electron
impact mass spectra (EI MS and HREI-MS) were recorded
on a Finnigan MAT 311 mass spectrometer (MASPEC Data
System). The 1H-NMR and 13C-NMR spectra were recorded
in deuterated DMSO on Avance Bruker AMX-300, AMX-
400 MHz, AMX-500 MHz machines. Chemical shifts (δ)
were shown on ppm scale, and coupling constants or J value
was expressed in Hz.
Conclusion
In vitro urease inhibitory activity was evaluated for synthetic
aryl hydrazones bearing thiazolyl coumarin hybrids 1–32. A
number of derivatives presented good to moderate inhibitory
potential and especially compound 21 was recognized as
potent urease inhibitor than the standard acetohydroxamic
acid. SAR proposed that analogs bearing -F, -Cl, -OH, and
-OMe substituents at particular positions on aryl ring were
found to be better active. Molecular docking studies revealed
noteworthy interactions between ligands (compounds) and
active site of urease enzyme. This study resulted in the rec-
ognition of leads for urease inhibition those could be used in
future research in order to identify drugs for urease-driven
ailments.
General procedure for the syntheses of aryl
hydrazones bearing thiazolyl coumarin
hybrids 1–32
Arylaldehyde (0.5 mmol) and thiosemicarbazide (0.5 mmol)
in ethanol (10 mL) were taken in round-bottomed fask. Few
drops of glacial acetic acid were added and refuxed for 2 h
with continuous stirring. Formation of aryl thiosemicar-
bazone derivative (intermediate) was monitored by TLC
analysis. After that, 3-bromoacetyl coumarin (0.5 mmol)
and triethylamine (0.75 mmol) were added into the reaction
mixture and further refuxed for 3 h. Precipitate appeared
in the reaction pot. Again TLC was monitored in order to
check the reaction completion. On complete disappearance
of intermediate, precipitates were fltered, washed with dis-
tilled water, and dried in vacuo. Crude product was crystal-
lized from ethyl acetate to aford further purifed compound.
Experimental
Materials and methods
Reagents and solvents were obtained from Sigma-Aldrich
(USA) and Merck (Germany). Thin layer chromatography
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Journal of the Iranian Chemical Society
rel. abund.): 411 (M+, 87), 413 (M+ + 2, 37), 376 (8),
270 (9), 244 (100), 211 (37), 172 (49), 145 (7), 128 (5);
HREI-MS calcd for C20H14ClN3O3S: m/z = 411.0444,
found 411.0440.
Spectral data of aryl hydrazones bearing
thiazolyl coumarin hybrids 1–32
(E)‑3‑(2‑(2‑Benzylidenehydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (1)
(E)‑2‑Methoxy‑4‑((2‑(4‑(2‑oxo‑2H‑chromen‑3‑yl)
thiazol‑2‑yl)hydrazono)methyl)phenyl acetate (4)
1
Yield: 78%; M.p.: 230–232 °C; H-NMR (300 MHz,
DMSO-d6): δ 12.18 (s, 1H, NH), 8.54 (s, 1H, H-5'), 8.06
(s, 1H, -N = CH), 7.86 (d, J5,6 = 7.8 Hz, 1H, H-5), 7.77
(s, 1H, H-4), 7.67 (m, 3H, H-7, H-3'', H-5''), 7.46 (m,
5H, H-6, H-8, H-2'', H-4'', H-6''); 13C-NMR (100 MHz,
DMSO-d6): δ 167.6, 158.7, 152.2, 143.9, 141.7, 138.1,
134.2, 131.6, 129.3, 128.8, 128.8, 126.2, 126.2, 124.6,
120.5, 119.1, 115.8, 110.6; EI MS m/z (% rel. abund.): 347
(M+, 82), 244 (100), 211 (31), 172 (41), 145 (7), 128 (5);
HREI-MS calcd for C19H13N3O2S: m/z = 347.0728, found
347.0724.
1
Yield: 76%; M.p.: 243–245 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.26 (s, 1H, NH), 8.54 (s, 1H, H-5'), 8.05
(s, 1H, -N = CH-), 7.86 (d, J5,6 = 7.6 Hz, 1H, H-5), 7.77
(s, 1H, H-4), 7.65 (t, J7,6 = J7,8 = 8.4 Hz, 1H, H-7), 7.46
(d, J8,7 =8.4 Hz, 1H, H-8), 7.41 (t, J6,5 =J6,7 =7.6 Hz, 2H,
H-6, H-2''), 7.28 (d, J5'',6'' = 8.4 Hz, 1H, H-5''), 7.15 (d,
J6'',5'' =8.0 Hz, 1H, H-6''), 3.83 (s, 3H, OCH3), 2.26 (s, 3H,
O = C-CH3); 13C-NMR (125 MHz, DMSO-d6): δ 168.3,
167.6, 158.7, 152.2, 151.1, 143.9, 140.9, 140.1, 138.1,
133.1, 131.6, 128.7, 124.6, 123.2, 120.5, 119.1, 118.8,
115.8, 110.6, 109.9, 55.7, 20.3; EI MS m/z (% rel. abund.):
435 (M+, 32), 393 (19), 244 (100), 211 (44), 172 (43), 149
(35), 102 (25), 51 (24); HREI-MS calcd for C22H17N3O5S:
m/z=435.0889, found 435.0879.
(E)‑3‑(2‑(2‑(3‑Methoxybenzylidene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (2)
1
Yield: 80%; M.p.: 256–258 °C; H-NMR (300 MHz,
(E)‑3‑(2‑(2‑(2‑Ethoxy‑3‑methoxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (5)
DMSO-d6): δ 12.20 (s, 1H, NH), 8.54 (s, 1H, H-5'), 8.03
(s, 1H, -N = CH), 7.86 (d, J5,6 = 6.3 Hz, 1H, H-5), 7.77
(s, 1H, H-4), 7.65 (t, J7,6 = J7,8 = 8.4 Hz, 1H, H-7), 7.46
(d, J8,7 = 8.4 Hz, 1H, H-8), 7.41 (t, J6,5 = J6,7 = 7.5 Hz,
1H, H-6), 7.37 (t, J5'',4'' = J5'',6'' = 7.8 Hz, 1H, H-5''),
7.25 (m, 2H, H-2'', H-4''), 6.97 (dd, J6'',2''/6'',4'' = 1.8 Hz,
J6'',5'' = 7.8 Hz, 1H, H-6''), 3.79 (s, 3H, OCH3); 13C-NMR
(75 MHz, DMSO-d6): δ 167.6, 159.5, 158.7, 152.3, 144.0,
141.5, 138.1, 135.7, 131.7, 129.9, 128.8, 124.7, 120.5,
119.1, 118.9, 115.8, 115.2, 111.1, 110.7, 55.1; EI MS
m/z (% rel. abund.): 377 (M+, 62), 244 (100), 211 (16),
172 (21), 145 (3); HREI-MS calcd for C20H15N3O3S:
m/z = 377.0834, found 377.0836.
1
Yield: 75%; M.p.: 215–217 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.05 (s, 1H, NH), 8.53 (s, 1H, H-5'), 7.97
(s, 1H, -N=CH), 7.85 (d, J5,6 =7.2 Hz, 1H, H-5), 7.74 (s,
1H, H-4), 7.62 (t, J7,6 = J7,8 = 7.6 Hz, 1H, H-7), 7.46 (d,
J8,7 =8.0 Hz, 1H, H-8), 7.40 (t, J6,5 =J6,7 =8.0 Hz, 1H, H-6),
7.25 (bd.s, 1H, H-2''), 7.17 (d, J6'',5'' = 8.4 Hz, 1H, H-6''),
7.01 (d, J5'',6'' =8.4 Hz, 1H, H-5''), 4.06 (q, 2H, -CH2-), 3.79
(s, 3H, OCH3), 1.37 (t, J2''',1a''' = 6.8, J2''',1b''' = 7.2 Hz, 3H,
OCH3); 13C-NMR (75 MHz, DMSO-d6): δ 167.7, 158.7,
152.2, 150.3, 148.2, 143.9, 141.9, 138.0, 131.6, 128.7, 127.0,
124.6, 120.5, 120.3, 119.1, 115.8, 111.8, 110.3, 109.3, 63.6,
55.5, 14.6; EI MS m/z (% rel. abund.): 421 (M+, 29), 244
(100), 211 (52), 172 (47), 149 (36), 102 (36); HREI-MS
calcd for C22H19N3O4S: m/z=421.1096, found 421.1106.
(E)‑3‑(2‑(2‑(2‑Chloro‑3‑methoxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (3)
1
(E)‑3‑(2‑(2‑(3‑Ethoxy‑2‑methoxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (6)
Yield: 76%; M.p.: 271–273 °C; H-NMR (300 MHz,
DMSO-d6): δ 12.44 (s, 1H, NH), 8.54 (s, 1H, H-5'),
8.45 (s, 1H, -N = CH), 7.87 (d, J5,6 = 7.6 Hz, 1H, H-5),
7.79 (s, 1H, H-4), 7.64 (t, J7,6 = J7,8 = 7.6 Hz, 1H, H-7),
7.53 (d, J8,7 = 8.0 Hz, 1H, H-8), 7.46 (d, J6'',5'' = 8.4 Hz,
1H, H-6''), 7.40 (t, J6,5 = J6,7 = 8.4 Hz, 1H, H-6), 7.38 (t,
J5'',4'' = J5'',6'' = 8.4 Hz, 1H, H-5''), 7.16 (d, J4'',5'' = 8.0 Hz,
1H, H-4''), 3.87 (s, 3H, OCH3); 13C-NMR (100 MHz,
DMSO-d6): δ 167.3, 158.6, 154.9, 152.2, 144.0, 138.2,
137.7, 132.6, 131.6, 128.8, 127.7, 124.6, 120.7, 120.4,
119.1, 117.6, 115.8, 112.8, 110.9, 56.2; EI MS m/z (%
1
Yield: 77%; M.p.: 210–212 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.06 (s, 1H, NH), 8.53 (s, 1H, H-5'), 7.98
(s, 1H, -N=CH), 7.85 (d, J5,6 =8.0 Hz, 1H, H-5), 7.74 (s,
1H, H-4), 7.64 (t, J7,6 = J7,8 = 8.0 Hz, 1H, H-7), 7.46 (d,
J8,7 =8.0 Hz, 1H, H-8), 7.40 (t, J6,5 =J6,7 =7.6 Hz, 1H, H-6),
7.26 (bd.s, 1H, H-2''), 7.16 (d, J6'',5'' =8.4 Hz, 1H, H-6''), 6.99
(d, J5'',6'' =8.4 Hz, 1H, H-5''), 4.07 (q, 2H, -CH2-), 3.80 (s,
3H, OCH3), 1.35 (t, J2''',1a'''/2''',1b''' =6.8 Hz, 3H, OCH3); 13C-
NMR (100 MHz, DMSO-d6): δ 167.7, 158.7, 152.3, 149.4,
1 3

Journal of the Iranian Chemical Society
149.1, 143.9, 142.0, 138.1, 131.6, 128.8, 126.9, 124.7,
120.5, 120.3, 119.2, 115.8, 112.6, 110.3, 108.5, 63.7, 55.3,
14.6; EI MS m/z (% rel. abund.): 421 (M+, 19), 244 (100),
211 (52), 172 (47), 134 (38), 102 (33); HREI-MS calcd for
C22H19N3O4S: m/z=421.1096, found 421.1104.
HREI-MS calcd for C21H17N3O4S: m/z = 407.0940, found
407.0957.
(E)‑3‑(2‑(2‑(3,4,5‑Trimethoxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (10)
(E)‑3‑(2‑(2‑(2‑Hydroxy‑4‑methoxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (7)
1
Yield: 78%; M.p.: 240–242 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.19 (s, 1H, NH), 8.53 (s, 1H, H-5'), 7.98
(s, 1H, -N=CH), 7.85 (d, J5,6 =7.6 Hz, 1H, H-5), 7.76 (s,
1H, H-4), 7.64 (t, J7,6 = J7,8 = 7.2 Hz, 1H, H-7), 7.46 (d,
J8,7 = 8.4 Hz, 1H, H-8),0.7.41 (t, J6,5 = J6,7 = 7.6 Hz, 1H,
H-6), 6.98 (s, 2H, H-2'', H-6''), 3.82 (s, 6H, 2OCH3), 3.69 (s,
3H, OCH3); 13C-NMR (75 MHz, DMSO-d6): δ 167.6, 158.7,
153.2, 153.2, 152.3, 143.9, 141.6, 138.7, 138.1, 131.6,
129.8, 128.8, 124.7, 120.5, 119.1, 115.8, 110.5, 103.6,
103.6, 60.1, 55.8; EI MS m/z (% rel. abund.): 437 (M+, 92),
244 (100), 211 (38), 193 (8), 172 (21), 145 (4); HREI-MS
calcd for C22H19N3O5S: m/z=437.1045, found 437.1057.
1
Yield: 72%; M.p.: 275–277 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.00 (s, 1H, NH), 10.21 (s, 1H, OH), 8.53
(s, 1H, H-5'), 8.26 (s, 1H, -N=CH-), 7.86 (d, J5,6 =7.6 Hz,
1H, H-5), 7.74 (s, 1H, H-4), 7.62 (t, J7,6 = J7,8 = 7.6 Hz,
1H, H-7), 7.54 (d, J8,7 = 8.8 Hz, 1H, H-8), 7.46 (d,
J6'',5'' = 8.4 Hz, 1H, H-6''), 7.40 (t, J6,5 = J6,7 = 7.6 Hz, 1H,
H-6), 6.51 (dd, J5'',3'' = 2.0 Hz, J5'',6'' = 6.8 Hz, 1H, H-5''),
6.45 (d, J3'',5'' =2.0 Hz, 1H, H-3'') 3.74 (s. 3H, OCH3); 13C-
NMR (100 MHz, DMSO-d6): δ 167.3, 161.4, 158.6, 157.5,
152.2, 144.0, 140.8, 138.1, 131.6, 128.7, 127.9, 124.6,
120.5, 119.1, 115.8, 112.9, 110.0, 106.4, 100.9, 55.1; EI
MS m/z (% rel. abund.): 393 (M+, 61), 376 (36), 244 (100),
211 (60), 172 (53), 136 (28), 108 (72); HREI-MS calcd for
C20H15N3O4S: m/z=393.0783, found 393.0782.
(E)‑3‑(2‑(2‑(4‑Chlorobenzylidene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (11)
1
Yield: 72%; M.p.: 280–282 °C; H-NMR (400 MHz,
(E)‑3‑(2‑(2‑(4‑Hydroxy‑3‑methoxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (8)
DMSO-d6): δ 12.28 (s, 1H, NH), 8.53 (s, 1H, H-5'), 8.05
(s, 1H, -N=CH), 7.86 (d, J5,6 =7.6 Hz, 1H, H-5), 7.78 (s,
1H, H-4), 7.69 (d, J2'',3''/6'',5'' =8.4 Hz, 2H, H-2'', H-6''), 7.64
(t, J7,6 =J7,8 =8.0 Hz, 1H, H-7), 7.50 (d, J3'',2''/5'',6'' =8.4 Hz,
2H, H-3'', H-5''), 7.46 (d, J8,7 = 8.4 Hz, 1H, H-8), 7.40 (t,
J6,5 =J6,7 =7.6 Hz, 1H, H-6); 13C-NMR (100 MHz, DMSO-
d6): δ 167.5, 158.7, 152.3, 144.0, 140.4, 138.2, 133.7, 133.2,
131.7, 128.9, 128.9, 128.8, 127.9, 127.9, 124.7, 120.5,
119.1, 115.9, 110.8; EI MS m/z (% rel. abund.): 381 (M+,
83), 383 (M+ + 2, 34), 270 (4), 244 (100), 211 (33), 172
(59); HRESI-MS calcd for C19H13ClN3O2S: m/z=382.0417
[M+H]+, found 382.0399; IR in KBr (cm−1); 3413.8 (N–H),
3240.2, 3138.0 (C–H), 1706.9 (C = O), 1571.9 (C = N),
1465.8, 1417.6, 1379.0 (C = C), 1296.1, 1261.4 (C–O),
657.7 (C–Cl).
1
Yield: 75%; M.p.: 271–273 °C; H-NMR (400 MHz,
DMSO-d6): δ 11.98 (s, 1H, NH), 9.43 (s, 1H, OH), 8.53 (s,
1H, H-5'), 7.95 (s, 1H, -N=CH-), 7.85 (d, J5,6 =7.6 Hz, 1H,
H-5), 7.73 (s, 1H, H-4), 7.64 (t, J7,6 =J7,8 =8.0 Hz, 1H, H-7),
7.45 (d, J8,7 =8.0 Hz, 1H, H-8), 7.40 (t, J6,5 =J6,7 =7.6 Hz,
1H, H-6), 7.07 (d, J6'',5'' = 8.4 Hz, 1H, H-6''), 6.82 (d,
J5'',6'' =8.0 Hz, 1H, H-5''), 3.81 (s, 3H, OCH3); EI MS m/z
(% rel. abund.): 393 (M+, 40), 244 (100), 211 (20), 172 (15),
145 (3); HREI-MS calcd for C20H15N3O4S: m/z=393.0783,
found 393.0796.
(E)‑3‑(2‑(2‑(3,4‑Dimethoxybenzylidene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (9)
1
Yield: 73%; M.p.: 205–207 °C; H-NMR (400 MHz,
(E)‑3‑(2‑(2‑(3‑Chlorobenzylidene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (12)
DMSO-d6): δ 12.18 (s, 1H, NH), 8.54 (s, 1H, H-5'), 8.32
(s, 1H, -N=CH), 7.87 (d, J5,6 =7.6 Hz, 1H, H-5), 7.77 (s,
1H, H-4), 7.64 (t, J7,6 = J7,8 = 8.4 Hz, 1H, H-7), 7.46 (d,
J8,7 =8.4 Hz, 1H, H-8), 7.40 (m, 2H, H-2'', H-6''), 7.14 (t,
J6,5 = J6,7 = 8.0 Hz, 1H, H-6), 7.07 (d, J5'',6'' = 7.6 Hz, 1H,
H-5''), 3.82 (s, 3H, OCH3), 3.77 (s, 3H, OCH3); 13C-NMR
(100 MHz, DMSO-d6): δ 167.6, 158.7, 152.6, 152.2, 147.2,
143.9, 138.1, 137.3, 131.6, 128.8, 127.6, 124.6, 124.3,
120.4, 119.1, 116.4, 115.8, 113.5, 110.6, 61.0, 55.7; EI MS
m/z (% rel. abund.): 407 (M+, 45), 376 (26), 361 (3), 259 (3),
244 (100), 211 (49), 172 (53), 149 (36), 102 (30), 77 (50);
1
Yield: 76%; M.p.: 130–132 °C; H-NMR (400 MHz,
DMSO-d6): δ 8.27 (s, 1H, H-5'), 8.04 (s, 1H, -N = CH),
7.68 (d, J4,5 =2.0 Hz, 1H, H-4), 6.98 (dd, J5,7/4'',2'' =1.2 Hz,
J5,6/4'',5'' =7.6 Hz, 2H, H-5, H-2''), 6.84 (t, J7,6/5'',4'' =8.4 Hz,
J7,8/5'',6'' = 8.8 Hz, 2H, H-7, H-5''), 6.61 (m, 4H, H-6, H-8,
H-2'', H-6''); EI MS m/z (% rel. abund.): 381 (M+, 11), 383
(M+ +2, 3), 314 (8), 285 (26), 270 (39), 257 (26), 244 (99),
229 (35), 172 (56), 156 (29), 139 (33); HRESI-MS calcd for
C19H13ClN3O2S: m/z=382.0417 [M+H]+, found 382.0407.
1 3

Journal of the Iranian Chemical Society
H-6), 7.25 (dd, J4'',6'' =2.8 Hz, J4'',5'' =6.0 Hz, 1H, H-4''), 6.92
(d, J3'',4'' =8.8 Hz, 1H, H-3''); 13C-NMR (100 MHz, DMSO-
d6): δ 167.3, 158.6, 154.5, 152.2, 144.0, 138.1, 137.4,
131.6, 129.8, 128.7, 124.6, 124.5, 123.2, 122.0, 120.4,
119.1, 117.9, 115.8, 110.6; EI MS m/z (% rel. abund.): 397
(M+, 50), 399 (M+ +2, 19), 244 (100), 230 (21), 211 (18),
172 (21), 155 (15); HRESI-MS calcd for C19H13ClN3O3S:
m/z=398.0366 [M+H]+, found 398.0370.
(E)‑3‑(2‑(2‑(2,4‑Dichlorobenzylidene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (13)
1
Yield: 80%; M.p.: 285–287 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.50 (s, 1H, NH), 8.54 (s, 1H, H-5'), 8.40
(s, 1H, -N = CH), 7.93 (d, J5,6 = 8.4 Hz, 1H, H-5), 7.87
(d, J6'',5'' = 6.8 Hz, 1H, H-6''), 7.81 (s, 1H, H-4), 7.69 (d,
J3'',5'' = 2.0 Hz, 1H, H-3''), 7.65 (t, J7,6 = J7,8 = 8.8 Hz, 1H,
H-7), 7.52 (dd, J5'',3'' = 2.0 Hz, J5'',6'' = 6.8 Hz, 1H, H-5''),
7.46 (d, J8,7 =8.0 Hz, 1H, H-8), 7.41 (t, J6,5 =J6,7 =7.6 Hz,
1H, H-6); EI MS m/z (% rel. abund.): 415 (M+, 85), 417
(M+ +2, 64), 380 (7), 244 (100), 211 (38), 172 (59), 145
(9); HRESI-MS calcd for C19H12Cl2N3O2S: m/z=416.0027
[M+H]+, found 416.0043.
(E)‑3‑(2‑(2‑(3,5‑Dichloro‑2‑hydroxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (17)
1
Yield: 76%; M.p.: above 300 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.51 (s, 1H, NH), 10.61 (s, 1H, OH), 8.54
(s, 1H, H-5'), 8.31 (s, 1H, -N=CH-), 7.86 (d, J5,6 =7.6 Hz,
1H, H-5), 7.81 (s, 1H, H-4), 7.65 (t, J7,6 = J7,8 = 7.2 Hz,
1H, H-7), 7.61 (d, J4'',6'' = 2.2 Hz, 1H, H-4''), 7.55 (d,
J6'',4'' =2.4 Hz, 1H, H-6''), 7.46 (d, J8,7 =8.0 Hz, 1H, H-8),
7.41 (t, J6,5 =J6,7 =7.6 Hz, 1H, H-6); 13C-NMR (125 MHz,
DMSO-d6): δ 158.7, 152.3, 138.9, 138.4, 131.8, 129.4,
128.9, 125.2, 124.7, 123.8, 122.3, 119.1, 115.9, 110.9; EI
MS m/z (% rel. abund.): 431 (M+, 39), 433 (M+ + 2, 29),
414 (33), 244 (100), 230 (13), 211 (16), 189 (20), 172 (19);
HRESI-MS calcd for C19H12Cl2N3O3S: m/z = 431.9976
[M+H]+, found 431.9991.
(E)‑3‑(2‑(2‑(3‑Chloro‑4‑hydroxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (14)
1
Yield: 73%; M.p.: 215–217 °C; H-NMR (400 MHz,
DMSO-d6): δ 8.49 (s, 1H, H-5'), 7.77 (s, 1H, -N = CH-),
7.67 (s, 1H, H-4), 7.62 (d, J5,6 = 7.4 Hz, 1H, H-5), 7.52
(bd.s, 1H, H-2''), 7.50 (t, J7,6 =J7,8 =7.4 Hz, 1H, H-7), 7.34
(d, J8,7 = 8.4 Hz, 1H, H-8), 7.29 (m, 2H, H-6, H-6''), 6.86
(d, J5'',6'' = 8.4 Hz, 1H, H-5''); EI MS m/z (% rel. abund.):
397 (M+, 19), 399 (M+ + 2, 7), 321 (2), 244 (100), 211
(56), 172 (62), 128 (21), 102 (26); HREI-MS calcd for
C19H12ClN3O3S: m/z=397.0288, found 397.0284.
(E)‑3‑(2‑(2‑(3‑Bromo‑5‑chlorobenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (18)
(E)‑3‑(2‑(2‑(4‑Chloro‑3‑hydroxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (15)
1
Yield: 72%; M.p.: above 300 °C; H-NMR (400 MHz,
DMSO-d6): δ 8.55 (s, 1H, H-5'), 8.29 (s, 1H, -N = CH-),
7.86 (d, J5,6 =8.0 Hz, 1H, H-5), 7.81 (bd.s, 1H, H-4), 7.65
(m, 4H, H-7, H-2'', H-4'', H-6''), 7.46 (d, J8,7 =8.4 Hz, 1H,
H-8),0.7.41 (t, J6,5 = J6,7 = 7.6 Hz, 1H, H-6); 13C-NMR
(125 MHz, DMSO-d6): δ 166.6, 158.7, 152.3, 151.5, 139.8,
138.4, 132.2, 131.8, 128.9, 126.5, 124.7, 124.2, 120.4,
119.1, 115.9, 111.8, 110.9; EI MS m/z (% rel. abund.): 458
(M+, 11), 460 (M+ +2, 14), 244 (100), 233 (26), 211 (18),
172 (16), 145 (8); HREI-MS calcd for C19H11BrClN3O2S:
m/z=457.9361, found 457.9364.
1
Yield: 74%; M.p.: 250–252 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.07 (s, 1H, NH), 10.59 (s, 1H, OH), 8.53
(s, 1H, H-5'), 7.94 (s, 1H, -N=CH-), 7.85 (d, J5,6 =7.6 Hz,
1H, H-5), 7.75 (s, 1H, H-4), 7.64 (t, J7,6 =J7,8 =7.6 Hz, 1H,
H-7), 7.62 (bd.s, 1H, H-2''), 7.46 (m, 2H, H-8, H-6''), 7.40
(t, J6,5 =J6,7 =7.6 Hz, 1H, H-6), 7.02 (d, J5'',6'' =8.4 Hz, 1H,
H-5''); 13C-NMR (100 MHz, DMSO-d6): δ 167.7, 158.7,
154.2, 152.2, 143.9, 140.6, 138.0, 131.6, 128.7, 127.5, 126.6,
126.3, 124.6, 120.5, 120.2, 119.1, 116.8, 115.8, 110.4; EI
MS m/z (% rel. abund.): 397 (M+, 32), 399 (M+ + 2, 22),
270 (5), 244 (100), 211 (40), 172 (45), 128 (24), 102 (24);
HREI-MS calcd for C19H12ClN3O3S: m/z=397.0254, found
397.0261.
(E)‑3‑(2‑(2‑(5‑Bromo‑2‑fuorobenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (19)
1
Yield: 71%; M.p.: 270–272 °C; H-NMR (400 MHz,
DMSO-d6): δ 8.54 (s, 1H, H-5'), 8.14 (s, 1H, -N = CH-),
7.92 (d, J6'',4'' =4.4 Hz, 1H, H-6''), 7.87 (d, J5,6 =8.0 Hz, 1H,
H-5), 7.79 (s, 1H, H-4), 7.64 (m, 2H, H-3'', H-4''), 7.46 (d,
J8,7 =8.4 Hz, 1H, H-8), 7.40 (t, J7,6 =J7,8 =7.6 Hz, 1H, H-7),
7.31 (t, J6,5 =J6,7 =9.6 Hz, 1H, H-6); 13C-NMR (100 MHz,
DMSO-d6): δ 167.2, 160.3, 158.7, 157.9, 152.3, 138.3,
133.4, 133.3, 132.7, 131.7, 128.8, 127.7, 124.7, 124.2,
124.1, 120.4, 119.1, 118.6, 118.4, 116.8, 115.9, 111.1; EI
(E)‑3‑(2‑(2‑(5‑Chloro‑2‑hydroxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (16)
1
Yield: 73%; M.p.: 280–282 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.24 (s, 1H, NH), 10.36 (s, 1H, OH), 8.53
(s, 1H, H-5'), 8.28 (s, 1H, -N=CH-), 7.86 (d, J5,6 =7.2 Hz,
1H, H-5), 7.77 (s, 1H, H-4), 7.64 (m, 2H, H-7, H-6''), 7.46
(d, J8,7 =8.0 Hz, 1H, H-8), 7.40 (t, J6,5 =J6,7 =7.6 Hz, 1H,
1 3

Journal of the Iranian Chemical Society
MS m/z (% rel. abund.): 443 (M+, 80), 445 (M+ + 2, 77),
425 (29), 270 (11), 244 (100), 211 (34), 199 (9), 172 (64),
145 (10), 128 (7); HREI-MS calcd for C19H11BrFN3O2S:
m/z=442.9739, found 442.9729.
270 (4), 244 (100), 211 (40), 172 (46), 145 (31), 102 (19);
HREI-MS calcd for C19H12BrN3O3S: m/z=440.9783, found
440.9774.
(E)‑3‑(2‑(2‑(3‑(Benzyloxy)‑4‑methoxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (23)
(E)‑3‑(2‑(2‑(2‑Bromo‑5‑fuorobenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (20)
1
Yield: 78%; M.p.: 239–241 °C; H-NMR (400 MHz,
1
Yield: 79%; M.p.: 280–282 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.04 (s, 1H, NH), 8.53 (s, 1H, H-5'),
7.95 (s, 1H, -N = CH), 7.85 (d, J5,6 = 7.6 Hz, 1H, H-5),
7.76 (s, 1H, H-4), 7.64 (t, J7,6 = J7,8 = 8.4 Hz, 1H, H-7),
7.49 (d, J6'',5'' = J5'',6'' = 7.6 Hz, 2H, H-5'', H-6''), 7.44 (d,
J2'',6'' = 4.0 Hz, 1H, H-2''), 7.41 (m, 5H, H-6, H-8, H-2''',
H-4''', H-6'''), 7.18 (d, J6'',5'' = 8.8 Hz, 1H, H-6''),7.03 (d,
J5'',6'' =8.4 Hz, 1H, H-5''), 5.15 (s, 2H, -CH2-), 3.80 (s, 3H,
OCH3); EI MS m/z (% rel. abund.): 483 (M+, 12), 333 (4),
244 (75), 211 (22), 172 (27), 145 (19), 91 (2), 65 (47);
HREI-MS calcd for C27H21N3O4S: m/z = 483.1253, found
483.1265.
DMSO-d6): δ 12.57 (s, 1H, NH), 8.55 (s, 1H, H-5'), 8.33
(s, 1H, -N=CH), 7.88 (d, J5,6 =6.8 Hz, 1H, H-5), 7.81 (s,
1H, H-4), 7.74 (m, 1H, H-6''), 7.65 (m, 2H, H-7, H-3''), 7.46
(d, J8,7 =8.4 Hz, 1H, H-8), 7.41 (t, J6,5 =J6,7 =7.6 Hz, 1H,
H-6), 7.24 (m, 1H, H-4''); 13C-NMR (125 MHz, DMSO-d6):
δ 167.2, 162.3, 160.4, 158.7, 152.3, 144.0, 138.5, 138.3,
135.1, 135.0, 134.9, 131.7, 128.8, 124.6, 120.4, 119.1, 118.2,
118.0, 117.0, 115.8, 112.5, 112.3, 111.2; EI MS m/z (% rel.
abund.): 443 (M+, 57), 445 (M+ +2, 64), 363 (16), 270 (8),
244 (100), 211 (27), 172 (57), 145 (7); HREI-MS calcd for
C19H11BrFN3O2S: m/z=442.9739, found 442.9729.
(E)‑3‑(2‑(2‑(4‑(Dimethylamino)benzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (24)
(E)‑3‑(2‑(2‑(5‑Fluoro‑2‑hydroxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (21)
1
Yield: 79%; M.p.: 275–277 °C; H-NMR (400 MHz,
1
Yield: 70%; M.p.: 295–297 °C; H-NMR (400 MHz,
DMSO-d6): δ 11.84 (s, 1H, NH), 8.52 (s, 1H, H-5'), 7.93
(s, 1H, -N=CH), 7.85 (d, J5,6 =7.2 Hz, 1H, H-5), 7.71 (s,
1H, H-4), 7.64 (t, J7,6 = J7,8 = 7.6 Hz, 1H, H-7), 7.48 (d,
J2''',3'''/6''',5''' =7.8 Hz, 2H, H-2''', H-6'''), 7.46 (d, J8,7 =10.4 Hz,
1H, H-8),0.7.40 (t, J6,5 = J6,7 = 7.6 Hz, 1H, H-6), 6.75 (d,
J3'',2''/5'',6'' =8.4 Hz, 2H, H-3'', H-5''), 2.95 (s, 6H, -N-(CH3)2);
13C-NMR (100 MHz, DMSO-d6): δ 167.8, 151.1, 142.8,
138.0, 131.6, 128.7, 127.6, 127.6, 124.7, 119.2, 115.8,
111.9, 111.9, 111.9, 110.0; EI MS m/z (% rel. abund.): 390
(M+, 35), 244 (100), 211 (38), 174 (24), 146 (45), 120 (24),
102 (24); HREI-MS calcd for C21H18N4O2S: m/z=390.1150,
found 390.1136.
DMSO-d6): δ 12.23 (s, 1H, NH), 10.04 (s, 1H, OH), 8.54 (s,
1H, H-5'), 8.31 (s, 1H, -N=CH-), 7.86 (d, J5,6 =6.8 Hz, 1H,
H-5), 7.77 (s, 1H, H-4), 7.64 (t, J7,6 =J7,8 =7.6 Hz, 1H, H-7),
7.46 (d, J8,7 =8.4 Hz, 1H, H-8), 7.40 (t, J6,5 =J6,7 =8.8 Hz,
1H, H-6), 7.36 (d, J6'',4'' =3.2 Hz, 1H, H-6''), 7.08.(m, 1H,
H-4''), 6.91 (m, 1H, H-3''); 13C-NMR (75 MHz, DMSO-
d6): δ 167.4, 158.7, 157.1, 154.0, 152.3, 152.1, 144.0,
138.2, 138.0, 131.7, 128.8, 124.7, 121.4, 121.3, 120.5,
119.1, 117.4, 117.3, 117.2, 116.9, 115.8, 111.0, 110.6; EI
MS m/z (% rel. abund.): 381 (M+, 81), 364 (100), 336 (5),
320 (5), 270 (7), 244 (91), 211 (60), 172 (73), 139 (27), 83
(58); HRESI-MS calcd for C19H13FN3O3S: m/z=382.0661
[M+H]+, found 382.0644.
(E)‑3‑(2‑(2‑(2,3,4‑Trihydroxybenzylidene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (25)
(E)‑3‑(2‑(2‑(3‑Bromo‑4‑hydroxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (22)
1
Yield: 70%; M.p.: 270–272 °C; H-NMR (400 MHz,
DMSO-d6): δ 11.98 (s, 1H, NH), 9.65 (s, 1H, OH), 9.46 (s,
1H, OH), 8.54 (s, 1H, H-5'), 8.50 (s, 1H, OH), 8.21 (s, 1H,
-N=CH-), 7.86 (d, J5,6 =8.0 Hz, 1H, H-5), 7.75 (s, 1H, H-4),
7.64 (t, J7,6 =J7,8 =8.4 Hz, 1H, H-7), 7.45 (d, J8,7 =8.0 Hz,
1H, H-8), 7.40 (t, J6,5 = J6,7 = 7.6 Hz, 1H, H-6), 6.90 (d,
J6'',5'' =8.4 Hz, 1H, H-6''), 6.39 (d, J5'',6'' =8.4 Hz, 1H, H-5'');
13C-NMR (75 MHz, DMSO-d6): δ 167.1, 158.7, 152.2,
148.0, 146.1, 143.2, 138.2, 132.7, 131.6, 128.7, 124.7,
120.4, 119.1, 118.4, 115.8, 111.9, 109.8, 107.8; EI MS m/z
(% rel. abund.): 395 (M+, 11), 244 (100), 216 (14), 174
(20), 153 (17), 102 (14); HREI-MS calcd for C19H13N3O5S:
m/z=395.0576, found 395.0589.
1
Yield: 74%; M.p.: 251–252 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.08 (s, 1H, NH), 10.67 (s, 1H, OH), 8.53
(s, 1H, H-5'), 7.94 (s, 1H, -N=CH-), 7.85 (d, J5,6 =7.2 Hz,
1H, H-5), 7.78 (d, J2'',6'' = 1.6 Hz, 1H, H-2''), 7.75 (s,
1H, H-4), 7.64 (t, J7,6 = J7,8 = 8.4 Hz, 1H, H-7), 7.50 ((d,
J6'',5'' =8.4 Hz, 1H, H-6''), 7.45 (d, J8,7 =8.0 Hz, 1H, H-8),
7.40 (t, J6,5 =J6,7 =7.6 Hz, 1H, H-6), 7.00 (d, J5'',6'' =8.4 Hz,
1H, H-5''); 13C-NMR (75 MHz, DMSO-d6): δ 167.7, 158.7,
155.2, 152.3, 143.9, 140.6, 138.1, 131.6, 130.5, 128.8,
127.0, 124.7, 120.5, 119.1, 116.6, 115.8, 110.4, 109.8; EI
MS m/z (% rel. abund.): 443 (M+, 21), 445 (M+ + 2, 20),
1 3

Journal of the Iranian Chemical Society
1H, H-4), 7.64 (t, J7,6 = J7,8 = 8.4 Hz, 1H, H-7), 7.49 (d,
J2''',3'''/6''',5''' = 7.6 Hz, 2H, H-2''', H-6'''), 7.41 (m, 4H, H-6,
H-3''', H-5''', H-4'''), 7.35 (d, J8,7 = 7.2 Hz, 1H, H-8), 7.29
(bd.s, 1H, H-2''), 7.16 (d, J6'',5'' =8.8 Hz, 1H, H-6''),7.10 (d,
J5'',6'' =8.4 Hz, 1H, H-5''), 5.12 (s, 2H, -CH2-), 3.82 (s, 3H,
OCH3); 13C-NMR (75 MHz, DMSO-d6): δ 167.7, 158.7,
152.3, 149.4, 149.1, 141.8, 138.1, 136.8, 131.6, 128.8,
128.4, 128.4, 127.9, 127.8, 127.8, 127.4, 124.7, 120.5,
120.2, 119.2, 115.8, 113.4, 110.3, 108,7, 69.9, 55.5; EI
MS m/z (% rel. abund.): 483 (M+, 6), 244 (86), 211 (16),
172 (9), 145 (5), 91 (100), 65 (30); HREI-MS calcd for
C27H21N3O4S: m/z=483.1253, found 483.1247.
(E)‑3‑(2‑(2‑(2‑Nitrobenzylidene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (26)
1
Yield: 73%; M.p.: 265–267 °C; H-NMR (400 MHz,
DMSO-d6): δ 8.55 (s, 1H, H-5'), 8.45 (s, 1H, -N=CH-), 8.06
(t, J7,6 =J7,8 =J5'',4'' =J5'',6'' =9.2 Hz, 2H, H-7, H-5''), 7.88 (d,
J5,6 =8.4 Hz, 1H, H-5), 7.80 (m, 2H, H-4, H-4''), 7.65 (m,
2H, H-3'', H-6''), 7.46 (d, J8,7 = 8.0 Hz, 1H, H-8), 7.41 (t,
J6,5 =J6,7 =7.6 Hz, 1H, H-6); EI MS m/z (% rel. abund.): 392
(M+, 45), 360 (10), 345 (17), 244 (100), 229 (16), 211 (16),
172 (42); HREI-MS calcd for C19H12N4O4S: m/z=392.0579,
found 392.0556.
(E)‑3‑(2‑(2‑((2‑Hydroxynaphthalen‑1‑yl)methylene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (30)
(E)‑3‑(2‑(2‑(4‑(Benzyloxy)benzylidene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (27)
1
1
Yield: 70%; M.p.: 272–274 °C; H-NMR (400 MHz,
Yield: 75%; M.p.: 223–225 °C; H-NMR (400 MHz,
DMSO-d6): δ 12.22 (s, 1H, NH), 10.90 (s, 1H, OH), 8.98
(s, 1H, H-5'), 8.76 (d, J5,6 =8.4 Hz, 1H, H-5), 8.58 (s, 1H,
-N = CH-), 7.87 (m, 4H, H-4, H-7'', H-5'', H-8''), 7.65 (t,
J7,6 =J7,8 =7.2 Hz, 1H, H-7), 7.60 (t, J6'',5'' =J6'',7'' =8.0 Hz,
1H, H-6''), 7.47 (d, J8,7 = 8.4 Hz, 1H, H-8), 7.41 (t,
J6,5 = J6,7 = 7.6 Hz, 1H, H-6), 7.38 (d, J4'',3'' = 6.8 Hz,
1H, H-4''), 7.23 (d, J3'',4'' = 8.8 Hz, 1H, H-3''); 13C-NMR
(100 MHz, DMSO-d6): δ 167.1, 158.7, 156.3, 152.3, 141.3,
138.3, 131.9, 131.7, 130.9, 128.8, 128.7, 128.1, 127.6,
124.6, 123.4, 123.0, 119.1, 118.1, 115.8, 110.2, 110.1; EI
MS m/z (% rel. abund.): 413 (M+, 30), 396 (46), 244 (100),
211 (23), 170 (37), 115 (40), 102 (22); HREI-MS calcd for
C23H15N3O3S: m/z=413.0834, found 413.0840.
DMSO-d6): δ 12.04 (s, 1H, NH), 8.53 (s, 1H, H-5'), 8.00
(s, 1H, -N=CH-), 7.85 (d, J5,6 =6.8 Hz, 1H, H-5), 7.74 (s,
1H, H-4), 7.62 (m, 3H, H-7, H-2''', H-6'''), 7.46 (m, 3H, H-8,
H-3''', H-5'''), 7.41 (m, 4H, H-6, H-3'', H-5'', H-4'''), 7.09
(d, J2'',3''/6'',5'' =8.4 Hz, 2H, H-2'', H-6''), 5.14 (s, 2H, -CH2-),
3.80 (s, 3H, OCH3); EI MS m/z (% rel. abund.): 453 (M+,
9), 244 (90), 211 (29), 172 (20), 145 (9), 91 (100), 65 (41);
HREI-MS calcd for C26H19N3O3S: m/z = 453.1147, found
453.1136.
(E)‑3‑(2‑(2‑(3‑(Benzyloxy)benzylidene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (28)
1
Yield: 75%; M.p.: 225–227 °C; H-NMR (400 MHz,
(E)‑3‑(2‑(2‑((1H‑Indol‑3‑yl)methylene)hydrazinyl)
thiazol‑4‑yl)‑2H‑chromen‑2‑one (31)
DMSO-d6): δ 12.21 (s, 1H, NH), 8.54 (s, 1H, H-5'), 8.01
(s, 1H, -N=CH), 7.86 (d, J5,6 =7.2 Hz, 1H, H-5), 7.78 (s,
1H, H-4), 7.64 (t, J7,6 = J7,8 = 7.6 Hz, 1H, H-7), 7.48 (d,
J2''',6''' =7.2 Hz, 2H, H-2''', H-6'''), 7.47 (d, J8,7 =11.2 Hz, 1H,
H-8), 7.42 (m, 3H, H-6, H-3''', H-5'''), 7.36 (m, 3H, H-5'',
H-4'''), 7.29 (bd.s, 1H, H-2''), 7.24 (d, J6'',5'' = 7.6 Hz, 1H,
H-6''), 7.04 (dd, J4'',2'' =2.0, J4'',5'' =6.4 Hz, 1H, H-4''), 5.15
(s, 2H, -CH2-); 13C-NMR (75 MHz, DMSO-d6): δ 167.7,
158.7, 158.6, 152.3, 144.0, 141.4, 138.1, 136.9, 135.7,
131.7, 129.9, 128.8, 128.4, 128.4, 127.8, 127.7, 127.7,
124.7, 120.5, 119.2, 119.1, 116.1, 115.8, 111.9, 110.7, 69.2;
EI MS m/z (% rel. abund.): 453 (M+, 31), 362 (10), 333 (13),
270 (5), 244 (80), 211 (29), 172 (34), 145 (23), 91 (100);
HREI-MS calcd for C26H19N3O3S: m/z = 453.1147, found
453.1128.
1
Yield: 75%; M.p.: 248–250 °C; H-NMR (300 MHz,
DMSO-d6): δ 11.79 (s, 1H, NH), 11.49 (s, 1H, NH), 8.55
(s, 1H, H-5'), 8.27 (s, 1H, -N=CH-), 8.23 (d, J5,6 =6.0 Hz,
1H, H-5), 8.86 (d, J7'',6'' = 7.6 Hz, 1H, H-7''), 7.77 (d,
J2'',CH = 2.7 Hz, 1H, H-2''), 7.75 (s, 1H, H-4), 7.65.(t,
J7,6 =J7,8 =8.1 Hz, 1H, H-7), 7.46 (m, 3H, H-6, H-8, H-4''),
7.21 (m, 2H, H-5'', H-6''); 13C-NMR (100 MHz, DMSO-d6):
δ 168.0, 158.7, 152.2, 143.9, 139.6, 137.9, 137.0, 131.5,
129.5, 128.7, 124.6, 123.9, 122.5, 121.5, 120.6, 120.4,
119.2, 115.8, 111.8, 111.5, 109.8; EI MS m/z (% rel. abund.):
386 (M+, 22), 244 (100), 211 (27), 172 (17), 142 (67), 89
(27); HREI-MS calcd for C21H14N4O2S: m/z = 386.0837,
found 386.0843.
(E)‑3‑(2‑(2‑(4‑(Benzyloxy)‑3‑methoxybenzylidene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (29)
(E)‑3‑(2‑(2‑((5‑Methylthiophen‑2‑yl)methylene)
hydrazinyl)thiazol‑4‑yl)‑2H‑chromen‑2‑one (32)
1
Yield: 71%; M.p.: 245–247 °C; H-NMR (400 MHz,
1
Yield: 76%; M.p.: 272–275 °C; H-NMR (300 MHz,
DMSO-d6): δ 12.07 (s, 1H, NH), 8.53 (s, 1H, H-5'), 7.98
(s, 1H, -N=CH), 7.85 (d, J5,6 =6.8 Hz, 1H, H-5), 7.75 (s,
DMSO-d6): δ 12.04 (s, 1H, NH), 8.52 (s, 1H, H-5'), 8.14
1 3

Journal of the Iranian Chemical Society
(s, 1H, -N=CH), 7.85 (d, J5,6 =8.1 Hz, 1H, H-5), 7.74 (s,
1H, H-4), 7.65 (t, J7,6 = J7,8 = 8.4 Hz, 1H, H-7), 7.45 (d,
J8,7 =8.1 Hz, 1H, H-8), 7.41 (t, J6,5 =J6,7 =8.1 Hz, 1H, H-6),
7.16 (d, J3'',4'' =3.3 Hz, 1H, H-3''), 6.79 (bd.d, J4'',3'' =3.0 Hz,
1H, H-4''), 2.45 (s, 3H, CH3); 13C-NMR (75 MHz, DMSO-
d6): δ 167.3, 158.7, 152.3, 143.9, 141.7, 138.1, 137.3, 136.8,
131.6, 129.6, 128.7, 126.2, 124.7, 120.5, 119.1, 115.8,
110.5, 15.26; EI MS m/z (% rel. abund.): 367 (M+, 26), 244
(100), 216 (15), 211 (37), 172 (31), 145 (8), 124 (23), 97
(18); HRESI-MS calcd for C18H14N3O2S2: m/z=368.0527
[M+H]+, found 368.0522.
using MMFF94 force feld. For validation of the software,
the co-crystalized ligands were extracted and re-docked
into the active sites using default docking protocol in MOE.
To evaluate the quality of cognate ligand, their root mean
square deviation (RMSD) values were obtained. The RMSD
between co-crystallized and re-docked conformation was
found to be 0.48 Å (Fig. 6). The reliable RMSD assures the
reliability of the MOE software to continue docking stud-
ies with synthetic compounds. For each ligand, 30 poses
were generated and saved in MOE database. Finally, dock-
ing results were manually visualized to understand the bind-
ing pattern of selected compounds within binding pocket of
proteins.
In vitro urease inhibitory assay
Supplementary Information The online version contains supplemen-
tary material available at https://doi.org/10.1007/s13738-021-02377-8.
Urease inhibitory potential of S-naproxen derivatives 1–39
was performed by using reported method [13]. Reaction
mixtures comprising of enzyme solution 25 µL and bufer
solution 55 µL containing urea 100 mM were incubated with
test compounds 5 µL (1 mM concentration) for 15 min at
30 °C in 96-well plates. Ammonia production was measured
by using the indophenols method in order to determine the
urease activity. Briefy, each of phenol reagent 45 µL (1%
w/v phenol and 0.005% w/v sodium nitroprusside) and alkali
reagent 70 µL (0.5% w/v NaOH and 0.1% active chloride
NaOCl) was added to each well. The increasing absorbance
was measured at 630 nm after 50 min using a microplate
reader (Molecular Device, USA). All steps were performed
thrice in a fnal volume of 200 µL. The results (change in
absorbance per min) were calculated by using SoftMax Pro
software (Molecular Device, USA). Assays were performed
at pH 8.2 (0.01 M K2HPO4.3H2O, 1 mM EDTA, and 0.01 M
LiCl). Acetohydroxamic acid was used as the standard inhib-
itor of urease. Percentage inhibitions were calculated from
the following formula:
Acknowledgements The authors are thankful to the Pakistan Acad-
emy of Sciences for providing fnancial support to Project No. (5-9/
PAS/440).
References
1. B. Krajewska, Ureases I. Functional, catalytic and kinetic proper-
ties: a review. J. Mol. Cat. B: Enz. 59, 9–21 (2009)
2. B. Krajewska, Ureases: roles, properties and catalysis.
Wiadomości Chemiczne 56, 223–253 (2002)
3. J.B. Sumner, The isolation and crystallization of the enzyme ure-
ase preliminary paper. J. Biol. Chem. 69, 435–441 (1926)
4. M.M. Ahari-Mostafavi, A. Sharif, M. Mirzaei, M. Amanlou,
Novel and versatile methodology for synthesis of β-aryl-β-
mercapto ketone derivatives as potential urease inhibitors. J. Iran.
Chem. Soc. 11, 1113–1119 (2014)
5. L. Asadi, K. Gholivand, K. Zare, Phosphorhydrazides as urease
and acetylcholinesterase inhibitors: biological evaluation and
QSAR study. J. Iran. Chem. Soc. 13, 1213–1223 (2016)
6. M.A. Lodhi, U. Ashiq, R.A. Jamal, K.M. Khan, Study of vana-
dium complexes as novel inhibitors of Bacillus pasteurii and
Canavalia ensiformis urease enzyme. J. Chem. Soc. Pak. 37,
549–558 (2015)
(
)
% Inhibition = 100 − ODtestwell∕ODcontrol x 100
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