Syntheses and biological activities of new hybrid molecules containing different heterocyclic moieties

Article abstract of DOI:10.1002/ardp.201300161

4-Aryl-5-(pyridin-3-yl)-4H-1,2,4-triazole-3-(thi)oles 5-7, obtained starting from nicotinic acid hydrazide were converted to the corresponding Mannich bases 12-24 by the reaction with several heterocyclic amines in the presence of formaldehyde. The synthesis of S-alkylated compounds 8-11 was performed from the reaction of the corresponding triazol-5-thioles with various alkyl halides. The condensation of carbo(thio)amides 2-4 with 4-chlorophenacyl bromide afforded the corresponding 1,3-thia(oxa)zol-2(3H)-ylidene]pyridine-3- carbohydrazides 25-27. 1,3-Thia(oxa)zolidine derivatives 28-30 were obtained from the cyclization reaction between compounds 2-4 and ethyl bromoacetate. All newly synthesized compounds were screened for their antimicrobial, antiurease, and antilipase activities. The biological activity studies revealed that all the compounds screened showed good or moderate antimicrobial, antiurease, and/or antilipase activity.

Full text of DOI:10.1002/ardp.201300161

Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
1
Full Paper
Syntheses and Biological Activities of New Hybrid Molecules
Containing Different Heterocyclic Moieties
Sule Ceylan1, Hakan Bektas2, Hacer Bayrak3, Neslihan Demirbas3, Sengul Alpay-Karaoglu4, and
Serdar Ülker4
1
Department of Forest Industrial Engineering, Artvin Çoruh University, Artvin, Turkey
Faculty of Arts and Sciences, Department of Chemistry, Giresun University, Giresun, Turkey
Department of Chemistry, Karadeniz Technical University, Trabzon, Turkey
Department of Biology, Recep Tayyib Erdogan University, Rize, Turkey
2
3
4
4-Aryl-5-(pyridin-3-yl)-4H-1,2,4-triazole-3-(thi)oles 5–7, obtained starting from nicotinic acid hydrazide
were converted to the corresponding Mannich bases 12–24 by the reaction with several heterocyclic
amines in the presence of formaldehyde. The synthesis of S-alkylated compounds 8–11 was performed
from the reaction of the corresponding triazol-5-thioles with various alkyl halides. The condensation of
carbo(thio)amides 2–4 with 4-chlorophenacyl bromide afforded the corresponding 1,3-thia(oxa)zol-2
(3H)-ylidene]pyridine-3-carbohydrazides 25–27. 1,3-Thia(oxa)zolidine derivatives 28–30 were obtained
from the cyclization reaction between compounds 2–4 and ethyl bromoacetate. All newly synthesized
compounds were screened for their antimicrobial, antiurease, and antilipase activities. The biological
activity studies revealed that all the compounds screened showed good or moderate antimicrobial,
antiurease, and/or antilipase activity.
Keywords: Biological activity / Nicotinic acid hydrazide / 1,3-Thia(oxa)zole / 1,3-Thia(oxa)zolidinone / 1,2,4-Triazole
Received: April 30, 2013; Revised: July 23, 2013; Accepted: July 26, 2013
DOI 10.1002/ardp.201300161
Introduction
Numerous reports have emphasized their anticancer and
antibacterial activities [9, 10–17]. 1,3-Thiazolidinone and
1,3-oxazolidinone derivatives, which are important groups
of heterocyclic compounds, have been the subject of extensive
study, and a number of reports have emphasized their use in
medicinal chemistry [18–20]. These classes of compounds
have been reported as novel inhibitors of the bacterial
enzyme [21]. It has also been reported that certain bi- and
polyheterocyclic compounds bearing 1,3,4-oxadiazole, 1,3,4-
thiadiazole, 1,3-oxazolidinone, 1,3-thiazolidinone, and 1,2,4-
triazole nucleus possess diverse biological activities [22–28].
During recent years, the microorganisms have gained
increasing resistance against the commonly used drugs, and
as a result of this event, the human population affected with
life-treating infectious diseases caused by multidrug-resistant
Gram-positive and Gram-negative pathogenic bacteria has
increased to an alarming level around the world. Another
resistant disease, tuberculosis, has become an infection
leading to mortality, partly due to poverty and inequity
and partly due to the HIV/AIDS pandemic, which greatly
increase the risk of infection proceeding to overt disease with
In the last decades, considerable evidence has been accumu-
lated on the significances of five-membered heterocycles in
the medicinal chemistry. For instance, triazole derivatives
have attracted attention as an important nucleus in the class
of chemotherapeutic agents due to their wide range of
therapeutic activities. Among the important representatives
of this class of agents, fluconazole and itraconazole display a
broad spectrum of antifungal activity and reduced toxicity
when compared with imidazole antifungals [1–7]. Other
important triazole derivatives, vorozole, letrozole, and
anastrozole known as aromatase inhibitors, have been used
for the treatment of breast cancer [8].
1,3,4-Oxadiazoles have been established as another member
of the privileged structural class in medicinal chemistry [1, 9].
Correspondence: Neslihan Demirbas, Department of Chemistry,
Karadeniz Technical University, 61080 Trabzon, Turkey.
E-mail: neslihan@ktu.edu.tr
Fax: þ90 4623253196
ß 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

2
S. Ceylan et al.
Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
an increasing rate worldwide [9, 29–32]. Therefore, it is crucial
to develop new, potent, fast-acting antimicrobial agents with
no side effects.
as therapeutic agents for curing obesity [58]. However, some
side effects including fecal incontinence, flatulence, and
steatorrhea have been reported for orlistat [59, 60]. The
compounds possessing antilipase activity can be the alterna-
tive to orlistat.
Motivated by these findings and in continuation of our
ongoing efforts endowed with the discovery of nitrogenated
heterocycles with potential chemotherapeutic activities, we
report here the synthesis and investigation of biological
activities of new 1,2,4-triazole derivatives incorporating
various heterocyclic rings responsible for biological activity
in a single structure.
Urease enzymes, which are found in a variety of plants,
algae, fungi, and bacteria, have been known to accelerate
hydrolysis of urea to ammonia gas at a reaction rate at least
1014 over the spontaneous reaction [33]. Medically, bacterial
ureases, reported as important virulence factors, play a role in
the pathogenesis of many clinical conditions such as
pyelonephritis, hepatic coma, peptic ulceration, the forma-
tion of injection-induced urinary stones, and stomach cancer.
The active site containing two nickel(II) atoms of the native
enzyme binds three water molecules and a hydroxide ion
bridged between two nickel ions [34]. In the course of Results and discussion
enzymatic reaction, urea replaces these three water molecules
and bridges the two metal ions. The surrounding by a
hydrogen-bonding network strongly activates the inert urea
molecule; it is subsequently attacked by the hydroxide ion,
forming a tetrahedral transition state. As a result, ammonia is
released from the active site followed by the negatively
charged carbamate [35]. The latter decomposes rapidly and
spontaneously, yielding a second molecule of ammonia. The
ammonia generated may disrupt several metabolic functions
in a large number of animal tissues and organs [35].
The ammonia produced by urease displays toxic effect on
various gastric cell lines. Furthermore, urease activity has
been proposed to damage the gastric epithelium via its
interaction with the immune system by stimulating an
oxidative burst in human neutrophils [36]. H2O2 generated in
this oxidative burst probably reacts with ammonia and
chloride to yield the toxic monochloramine [33]. Finally, the
ammonia may reach the serum and contribute to symptoms
of hepatic encephalopathy in patients suffering from
cirrhosis. Apart from ammonia, the carbon dioxide generated
by urea hydrolysis may play a significant role in the survival of
Helicobacter pylori in the gastric mucosa [37–43].
Several classes of compounds have been reported as
antiurease agents, such as hydroxamic acids [35, 44, 45],
phosphoramidates [46, 47], polyphenols [48, 49], 1,2,4-
triazoles, 1,3,4-oxadiazoles, and 1,3,4-thiadiazoles [50–52].
Furthermore, some Schiff base-metal complexes have been
found to possess urease inhibitory effects [53, 54], along with
other metal complexes [55]. However, the presence of heavy-
metal atoms has restricted their applications as drugs in the
human body [56].
Obesity has been widely recognized as a major global health
problem, which is caused by a chronic imbalance between
energy intake and energy expenditure. Obesity can be the
reason for different serious diseases, including hypertension,
hyperlipidemia, arteriosclerosis, and type II diabetes [57].
Pancreatic lipase plays an important role in fat digestion.
Pancreatic lipase inhibitors, such as orlistat, have been used
Chemistry
The main aim of the present study is to design and synthesize
new bi- and polyheterocyclic compounds incorporating 1,2,4-
triazole, piperazine, morpholine, 1,3-thiazol(idinone), and/or
1,3-oxazol(idinone) ring within a single structure as pharma-
cophore groups and investigate their antimicrobial, antiur-
ease, and antilipase activities. This idea emerged from the
finding that the compounds obtained in our previous study
starting from isonicotinic acid hydrazide as hybrid molecules
containing several heterocyclic rings possess good to moder-
ate antimicrobial and/or antiurease activities [61, 62]. In
addition, several 1,2,4-triazoles, 1,3,4-oxadiazoles, and 1,3,4-
thiadiazole derivatives were reported to have antiurease
activity [50–52]. Moreover, different types of hybrid com-
pounds incorporating also 1,2,4-triazole nucleus displayed
antimicrobial and/or antilipase activity in one of our studies
(unpublished data). Furthermore, several Mannich bases of
triazole derivatives including piperazine, thiomorpholine, or
morpholine moiety were synthesized as antimicrobial agents
in our laboratory [63–65]. Synthesis of the intermediate and
target compounds was performed according to the reactions
outlined in Schemes 1 and 2. The substituents on compounds
2–30 are presented in Table 1.
The synthesis of carbo(thio)amides (2–4) was performed
from the reaction of nicotinic acid hydrazide with several
iso(thio)cyanates in ethanolic solution in good yields. The
FT-IR spectra of compounds 2, 4 displayed additional
absorption bands at 1195 and 1156 cmꢀ1 pointing to C S
–
–
stretching. In the 1H NMR spectra of compounds 2–4, the
signal due to amine function was not present; instead, new
signals that originated from hydrazide structure were
recorded at 7.45–10.67 ppm as D2O exchangeable peaks.
Moreover, additional signals belonging to benzyl or phenyl
moiety appeared at the related chemical shift values.
The intramolecular cyclization of compounds 2–4 in basic
media generated the corresponding 4-benzyl-5-pyridin-3-yl-4H-
1,2,4-triazole compounds (5–7). In the 1H NMR spectra of
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Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
Hybrid Molecules Containing Different Heterocyclic Moieties
3
R'
N
N
N
X
N
R
_
12 24
HCHO,
R''H
N
O
N
N
N
O
RNCX
X
NaOH
N
XH
NHNH2
N
NHNH
1
R
NHR
_
5 7
_
2 4
CH3I
CH3(CH2)5Cl
N
N
N N
N
S(CH2)5CH3
N
SCH3
N
R
N
R
10, 11
8, 9
Scheme 1. Synthetic pathway for the preparation of compounds 2–24.
Cl
N
O
X
NHN
N
N
O
R
X
NHNH
_
25 27
NHR
_
2 4
N
O
O
X
NHN
N
R
_
28 30
Scheme 2. Synthesis of compounds 25–30.
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Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
Table 1. Substituents on compounds 2–30.
compounds 5–7, the signal observed at 14.30 ppm was
attributed to –SH proton. The signal due to this group was
recorded at 2725–2763 cmꢀ1 in the FT-IR spectra. Moreover,
compounds 5–7 gave stable [Mþ1]þ, [MþNa]þ, and [MþK]þ
peaks in the mass spectra and also the elemental analysis data
are consistent with the assigned structures.
The synthesis of compounds 8–11 was performed by the
reaction of compounds 5–7 with the corresponding alkyl
halides in the presence of sodium ethoxide. With the
conversion of compounds 5–7 into the alkylated derivatives
(8–11), the signal that originated from –SH group disappeared
in the 1H and FT-IR spectra; instead new signals due to S-alkyl
moiety were observed at the related chemical shift values.
[M]þ or [MþNa]þ peaks were present in the mass spectra of
compounds 8–11 at the 305.26, 269.15, 339.30, and 345.31 m/z
values, respectively. Furthermore, these compounds dis-
played good elemental analysis results.
Comp. no.
R
R0
X
2
3
4
5
6
7
8
9
–CH2C6H5
–CH2C6H5
–C6H5
–CH2C6H5
–CH2C6H5
–C6H5
–CH2C6H5
–C6H5
–C6H5
–
–
–
–
–
–
–
–
–
S
O
S
S
O
S
S
S
S
S
10
11
–C6H5
12
–CH2C6H5
S
N
N
O
S
13
14
–CH2C6H5
–CH2C6H5
O
S
N
N
F
The Mannich base derivatives (12–24) were obtained by the
reaction of compounds 5–7 with several heterocyclic amines,
including morpholine, thiomorpholine, imidazole, or furan
nucleus, under mild reaction conditions in the presence of
formaldehyde solution. In the FT-IR spectra of these
compounds, no signal derived from SH or OH function was
present. The 1H NMR and 13C NMR spectra of compounds
12–24 exhibited additional signals due to amine moiety at the
related chemical shift values. Compounds 12–24 gave
elemental analysis results consistent with the proposed
structures. Furthermore, molecular masses of these com-
pounds were confirmed by the appearance of [M]þ, [Mþ1]þ,
[Mþ2]þ, or [MþNa]þ ion peak at corresponding m/z values in
the mass spectra of these compounds.
The synthesis of compounds 25–27 was performed from the
condensation between compounds 2–4 and 4-chlorophenacyl
bromide in ethanolic solution, while the treatment of the
same intermediates with ethyl bromoacetate afforded com-
pounds 28–30. The structures of compounds 25–30 were
elucidated on the basis of spectroscopic techniques such as 1H
NMR, 13C NMR, FT-IR, EI-MS, and elemental analysis.
NH (CH2)3
N
15
–CH2C6H5
S
N
N
N
O
S
16
17
–CH2C6H5
–CH2C6H5
O
O
N
N
N
N
F
18
19
–CH2C6H5
–C6H5
O
S
O
S
20
21
–C6H5
–C6H5
S
S
N
N
F
NH (CH2)3
Biological activity
N
22
23
–C6H5
–C6H5
S
S
All the newly synthesized compounds were screened for their
antimicrobial activities and the results obtained are presented
in Table 2. Compounds 2–9 displayed activity against some of
the test microorganisms with the MIC values varying between
7.8 and 500 mg/mL. Among these, compounds 4, 7, and 9 are
the most active ones toward Arthrobacter oxydans with the MIC
values 7.8, 15.6, and 31.3 mg/mL, respectively. Arthrobacter spp.,
being basic soil bacteria, were found to perform several
important functions. It was reported that several species of
Arthrobacter can reduce hexavalent chromium, which can
cause severe irritation in humans, and they are also known
to degrade agricultural pesticides. Among the S-alkylated
N
–NH(CH2)4CH3
NH CH2
24
25
–C6H5
S
S
O
–CH2C6H5
–
26
27
28
29
30
–CH2C6H5
–C6H5
–CH2C6H5
–CH2C6H5
–C6H5
–
–
–
–
–
O
S
S
O
S
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Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
Hybrid Molecules Containing Different Heterocyclic Moieties
5
Table 2. Antimicrobial activity of the compounds (mg/mL).
Microorganismsa and minimal inhibition concentration (MIC)
Comp. no
Ec.
Yp.
Pa.
Sa.
Ef.
Bc.
Ms.
Ca.
Sc.
Ko.
Ar.
Ct.
Pv.
Sm.
Ac.
2
3
4
5
6
7
8
9
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
250
–
125
125
7.8
250
–
–
–
–
–
–
–
–
–
–
250
–
250
–
500
–
–
500
–
250
–
250
–
250
125
–
125
–
–
500
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
250
15.6
–
–
–
–
–
–
–
–
–
–
–
–
–
250
–
–
–
–
–
–
–
–
–
–
–
–
–
–
125
–
–
–
–
–
–
–
–
–
–
–
–
–
–
31.3
<3.8
<3.8
7.8
7.8
–
–
–
–
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
Amp.
Strep
Flu.
15.6
62.5
31.3
31.3
500
31.3
31.3
31.3
–
15.6
62.5
31.3
31.3
500
31.3
31.3
31.3
–
15.6
62.5
31.3
31.3
500
31.3
62.5
31.3
–
<1.9
<1.9
31.3
31.3
250
31.3
31.3
31.3
–
<1.9
<1.9
15.6
15.6
250
15.6
31.3
31.3
–
7.8
<1.9
31.3
31.3
250
31.3
62.5
62.5
–
1.9
1.9
31.3
31.3
125
31.3
15.6
15.6
–
3.9
125
500
500
500
62.5
500
–
1.9
125
125
500
500
31.3
250
–
–
–
62.5
62.5
31.3
<3.9
–
0.9
1.9
250
125
–
–
–
–
7.8
125
125
–
250
62.5
–
125
125
125
125
250
250
250
–
62.5
–
7.8
15.6
7.8
15.6
7.8
7.8
31.3
31.3
15.6
15.6
125
125
7.8
250
125
7.8
<3.9
15.6
31.3
15.6
125
–
62.5
125
–
–
–
–
–
–
–
–
125
–
–
500
250
250
31.3
–
500
250
250
31.3
–
500
500
500
62.5
–
–
–
500
250
500
31.3
–
31.3
31.3
62.6
31.3
–
–
–
250
125
250
500
–
500
–
500
500
31.3
–
500
500
15.6
–
–
–
250
–
–
–
–
125
62.5
–
250
125
500
15.6
250
–
250
15.6
–
250
–
62.5
–
125
–
31.3
250
62.5
–
31.3
250
62.5
–
31.3
–
31.3
250
62.5
–
7.8
–
31.3
250
62.5
–
31.3
125
15.6
–
–
250
–
31.3
250
250
250
–
125
125
125
–
15.6
–
15.6
–
–
62.5
–
62.5
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
250
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
250
32
–
–
–
–
10
32
>128
35
10
15
–
–
32
32
>128
>128
35
>25
Nd.
Nd.
<8
Nd.
Nd.
Nd.
a
Ec., Escherichia coli ATCC 25922; Yp., Yersinia pseudotuberculosis ATCC 911; Pa., Pseudomonas aeruginosa ATCC 43288; Sa., Staphylococcus aureus
ATCC 25923; Ef., Enterococcus faecalis ATCC 29212; Bc., Bacillus cereus 702 Roma; Ms., M. smegmatis ATCC607; Ca., Candida albicans ATCC
60193; Sc., Saccharomyces cerevisiae RSKK 251; Ko., Klebsiella oxitoka (laboratory strain); Ar., Arthrobacter oxydans (laboratory strain); Ct.,
Candida tropicalis ATCC 13803; Pv., Proteus vulgaris ATCC 13315; Sm., Serretia marcescens (laboratory strain); Ac., Acinetobacter sp. (laboratory
strain); Amp., ampicillin; Strep., streptomycin; Flu., fluconazole; —, no activity; Nd., not detected.
triazoles (8–11), compounds 10 and 11 were found to be more
active on the test microorganisms. It can be concluded that
the increase in lipophilicity of compounds 10 and 11 due to
the presence of S-alkyl moiety leads to more active
compounds. Generally, Mannich bases (12–24) containing
several lipophilic substituents at position 2 of the 1,2,4-
triazole skeleton exhibited good antimicrobial activities
against the test microorganisms. Among these, the activities
of compounds 18 and 23 were more specific. Compound
18 that includes the 4-fluorophenylpiperazine nucleus at
position 2 of 1,2,4-triazole ring displayed excellent activities
on A. oxydans, Proteus vulgaris – a rod-shaped, Gram-negative
bacterium that inhabits the intestinal tracts of humans and
animals and is found in soil, water, and fecal matter, and
Serretia marcescens – a Gram-negative, rod-shaped bacterium
in the family Enterobacteriaceae, involved in nosocomial
infections, particularly catheter-associated bacteremia, uri-
nary tract infections, and wound infections. Other Mannich
base possessing specific activity (23) showed excellent activity
toward A. oxydans with the MIC value 15.6 mg/mL. Further,
this compound (23) was found to have slight activity on
S. marcescens. Compounds 25 and 26, which have a N0-[3-benzyl-
4-(4-chlorophenyl)-1,3-oxa(thia)zol-2(3H)-ylidene]pyridine-3-
carbohydrazide structure, exhibited good to moderate
activities against most of the test microorganisms with the
MIC values varying between 250 and 15.6 mg/mL. Another
1,3-thiazol-2(3H)-ylidene]pyridine-3-carbohydrazide derivative
(27) carrying a phenyl substituent instead of benzyl was found
to be quite active on A. oxydans with the MIC value 7.8 mg/mL.
This compound displayed slight activity against Klebsiella
oxitoka, a Gram-negative bacterium. Similarly, 5-oxo-1,3-
oxa(thia)zolidin-2-ylidene]pyridine-3-carbohydrazides (28–30)
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S. Ceylan et al.
Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
Table 3. Inhibitory activities of the synthesized compounds
against urease.
39.5 ꢁ 8.6 mg/mL was used as standard inhibitor. Potent
compounds have their IC50 values in the range of 23.2–
102.3 mg/mL. Among the synthesized compounds, compound
17 that has a thiomorpholine nucleus in the position 2 of the
5-pyridine-3-yl-1,2,4-triazol-3-one scaffold displayed the best
inhibitory effect against urease with an IC50 value of
23.2 ꢁ 4 mg/mL. Besides compound 17, also compounds 14,
15, 18, and 22 were determined to be more potent than the
standard, thiourea. Dose-dependent urease inhibitory effects
of compounds are depicted in Fig. 1.
The synthesized compounds were evaluated with regard to
pancreatic lipase activity and the obtained results are
presented in Table 4. Among all the compounds, compound
26 that has 1,3-oxazol-2(3H)-ylidene]pyridine-3-carbohydra-
zide core structure exhibited antilipase activities at various
concentrations. The compound inhibited pancreatic lipase
activity by 21.1, 74.6, and 98.2% at a concentration of 0.69,
2.08, and 6.25 mg/mL, respectively. Orlistat, a known pancre-
atic lipase inhibitor used as anti-obesity drug, showed
inhibitory effect by 71.5, 98.6, and 99.1% at the same
concentrations. IC50 values of orlistat and compound 26
were calculated as 0.42 and 0.87 mg/mL, respectively. Orlistat
is the only approved anti-obesity medication [58]. However, it
Residual
activity ꢁ S.D. (%)
IC50 ꢁ S.D.
(mg/mL)
Compound
2
4
13
14
15
16
17
18
99.0 ꢁ 9
74.9 ꢁ 14
69.5 ꢁ 1
66.5 ꢁ 4
46.1 ꢁ 4
90.1 ꢁ 15
37.7 ꢁ 12
70.2 ꢁ 2
58.6 ꢁ 12
86.2 ꢁ 1
102.3 ꢁ 4.7
63.2 ꢁ 32.1
44.9 ꢁ 1.6
38.5 ꢁ 5
28.2 ꢁ 3.6
59.6 ꢁ 9.1
23.2 ꢁ 4
40.8 ꢁ 1.8
40.0 ꢁ 5.7
39.5 ꢁ 8.6
22
Thiourea
All compounds and thiourea were assayed at a final concentra-
tion of 28 mg/mL.
exhibited specific activity toward K. oxitoka, A. oxydans, and/or
Candida tropicalis, yeast-like fungi. Among compounds 28–30,
compound 30 displayed maximum activity against A. oxydans
with the MIC value 7.8 mg/mL.
The synthesized compounds were assayed for their in vitro
inhibitory activity against Jack bean urease. Only positive
results are presented in Table 3. Thiourea with IC50 value
120
110
100
90
80
70
60
50
40
30
20
10
0
Thiourea
2
4
13
14
15
16
17
18
22
-10
0
50
100
150
200
250
300
Conc. (µg/mL)
Figure 1. Dose-dependent urease inhibitory effects of some of the synthesized compounds. Thiourea was used as standard inhibitor.
Inhibitory effects of all compounds and thiourea were measured at concentrations of 250–0.114 mg/mL. Residual activities of compounds are
expressed as the mean ꢁ S.D. in triplicate.
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Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
Hybrid Molecules Containing Different Heterocyclic Moieties
7
Table 4. Residual pancreatic lipase activity of synthesized
compounds.
DMSO-d6 on a Bruker Avance II 400 MHz NMR spectrometer
1
(400.13 MHz for H and 100.62 MHz for 13C). The chemical shifts
are given in ppm relative to Me4Si as an internal reference;
J values are given in Hz. The elemental analysis was performed on
a Costech Elemental Combustion System CHNS-O elemental
analyzer. All the compounds gave C, H, and N analyses within
ꢁ0.4% of the theoretical values. The mass spectra were obtained
on a Quattro LC–MS (70 eV) instrument. Compounds 2, 4, 5, and
6 are available commercially.
Compound
Residual activity (%)
Positive control
2
3
4
5
6
100
105
100
111
104
88
General method for the synthesis of compounds 2–4
A mixture of compound 1 (10 mmol) and the corresponding
isothiocyanate (10 mmol) was heated under reflux in ethanol for
3 h (the progress of the reaction was monitored by TLC). Then, the
mixture was kept overnight in the cold. The resulting solid
separated was collected by filtration and recrystallized from
dimethyl sulfoxide/water (1:3; for 2 and 4) or ethanol (for 3) to
afford the desired product.
7
8
9
107
107
81
80
64
81
67
62
85
64
36
42
101
87
95
102
95
102
76
2
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
N-Benzyl-2-(pyridin-3-ylcarbonyl)hydrazinecarbothio-
amide (2)
Yield: 73%, m.p. 195–196°C. IR (KBr, n, cmꢀ1): 3297 (3NH), 1658
(C O), 1195 (C S). 1H NMR (DMSO-d6, d ppm): 4.75 (s, 2H, CH2),
–
–
–
–
7.24 (d, 5H, arH, J ¼ 3.2 Hz), 7.51–7.57 (m, 1H, arH), 8.25 (d, 1H,
arH, J ¼ 7.8 Hz), 8.75 (s, 2H, arH þ NH), 9.08 (s, 1H, arH), 9.59 (s, 1H,
NH), 10.67 (s, 1H, NH). 13C NMR (DMSO-d6, d ppm): 47.47 (CH2), arC:
[124.15 (2CH), 127.34 (2CH), 127.73 (CH), 128.76 (CH), 128.91 (C),
–
136.30 (CH), 140.03 (C), 149.60 (CH), 153.06 (CH)], 165.47 (C O),
–
þ
þ
–
–
182.97 (C S). EI MS m/z (%): 287.27 ([Mþ1] , 36), 309.28 ([MþNa] ,
66
79
74
84
15), 180.03 (27), 166.13 (25), 138.02 (100), 132.80 (94), 121.04 (51).
Elemental analysis, for C14H14N4OS, calcd. (%): 58.72 C, 4.93 H,
19.57 N; Found (%): 59.08 C, 4.89 H, 19.40 N.
N-Benzyl-2-(pyridin-3-ylcarbonyl)hydrazinecarbox-
All compounds were tested at a final concentration of 6.25 mg/mL
and evaluated by comparing with positive control.
amide (3)
Yield: 70%, m.p. 164–165°C. IR (KBr, n, cmꢀ1): 3331 and 3219
(3NH), 1688 and 1671 (2C O). 1H NMR (DMSO-d6, d ppm): 4.25
–
–
has some side effects, such as fecal incontinence, flatulence,
and steatorrhea [59, 60]. Compound 26 can be considered a
good alternative to orlistat. Compounds 17 and 18 showed
moderate antilipase activity, which are Mannich bases of
1,2,4-triazol-3-one compounds containing 4-fluorophenylpi-
perazine (18) or thiomorpholine nucleus in their structures.
Dose-dependent pancreatic lipase activity is shown in Fig. 2.
(d, 2H, CH2, J ¼ 5.6 Hz), 7.21–7.28 (m, 6H, arH þ NH), 7.53 (t, 1H,
arH, J ¼ 5.4 Hz), 8.08 (s, 1H, NH), 8.23 (d, 1H, arH, J ¼ 8.0 Hz), 8.73
(d, 1H, arH, J ¼ 4.4 Hz), 9.06 (s, 1H, arH), 10.38 (s, 1H, NH). 13C NMR
(DMSO-d6, d ppm): 43.29 (CH2), arC: [124.19 (CH), 127.22 (CH),
127.60 (2CH), 128.81 (2CH), 129.09 (C), 136.07 (CH), 141.26 (C),
–
–
149.35 (CH), 152.95 (CH)], 159.08 (C O), 165.84 (C O). EI MS m/z
–
–
(%): 271.17 ([Mþ1]þ, 34), 293.24 ([MþNa]þ, 31), 214.18 (26), 192.99
(26), 181.04 (26), 166.01 (36), 136.87 (23), 149.97 (100), 137.89 (44),
106.95 (29). Elemental analysis, for C14H14N4O2, calcd. (%): 62.21
C, 5.22 H, 20.73 N; Found (%): 62.55 C, 5.60 H, 20.95 N.
Experimental
N-Phenyl-2-(pyridin-3-ylcarbonyl)hydrazinecarbothio-
General
amide (4)
All the chemicals were purchased from Fluka Chemie AG Buchs
(Switzerland) and used without further purification. Melting
points of the synthesized compounds were determined in open
capillaries on a Büchi B-540 melting point apparatus and are
uncorrected. Reactions were monitored by thin-layer chromatog-
raphy (TLC) on silica gel 60 F254 aluminium sheets. The mobile
phase was ethyl acetate/diethyl ether 1:1 and detection was
made using UV light. FT-IR spectra were recorded as potassium
Yield: 78%, m.p. 181–183°C. IR (KBr, n, cmꢀ1): 3219, 3163, and
1
–
–
3106 (3NH), 1682 (C O), 1156 (C S). H NMR (DMSO-d , d ppm):
–
–
6
7.45 (d, 7H, arH þ NH, J ¼ 14.0 Hz), 7.66 (d, 2H, arH þ NH,
J ¼ 7.8 Hz), 8.50 (s, 2H, arH þ NH), 8.59 (s, 1H, arH). 13C NMR
(DMSO-d6, d ppm): arC: [122.99 (C), 124.17 (CH), 129.47 (CH), 130.12
(2CH), 130.32 (2CH), 134.84 (C), 136.56 (CH), 149.37 (CH), 151.69
þ
–
–
–
–
(CH)], 149.21 (C O), 169.46 (C S). EI MS m/z (%): 273.12 ([Mþ1] ,
57), 301.17 (38), 188.90 (25), 148.84 (82), 137.83 (100), 120.85 (76),
104.87 (37). Elemental analysis, for C13H12N4OS, calcd. (%):
57.34 C, 4.44 H, 20.57 N; Found (%): 57.44 C, 4.58 H, 20.27 N.
bromide pellets using
a Perkin Elmer 1600 series FTIR
spectrometer. 1H NMR and 13C NMR spectra were registered in
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8
S. Ceylan et al.
Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
110
26
Orlistat
100
90
80
70
60
50
40
30
20
10
0
-10
0
1
2
3
4
5
6
Concentration (µg/mL)
Figure 2. Dose-dependent inhibition of pancreatic lipase by compound 26. Orlistat was used as standard inhibitor. Inhibitory effects of
compound 26 and orlistat were measured at concentrations of 6.25–0.003 mg/mL. Residual activities of compounds are expressed as the
mean ꢁ S.D. in triplicate.
General method for the preparation of compounds 5–7
A solution of corresponding carbothioamide 2–4 (10 mmol) in
ethanol/water (1:1) was refluxed in the presence of 2 N NaOH for
6 h (the progress of the reaction was monitored by TLC). Then, the
resulting solution was cooled to room temperature and acidified
to pH 5 with 37% HCl. The precipitate formed was filtered off,
washed with water, and recrystallized from ethanol/water (1:3)
(for 5), ethyl acetate (for 6), or dimethyl sulfoxide/water (1:3; for 7)
to afford the desired compound.
7.06 (s, 2H, arH), 7.24 (s, 3H, arH), 7.47 (d, 1H, arH, J ¼ 4.2 Hz), 7.92
(d, 1H, arH, J ¼ 7.8 Hz), 8.66 (d, 2H, arH, J ¼ 7.8 Hz), 12.32 (s, 1H,
NH). 13C NMR (DMSO-d6, d ppm): 44.79 (CH2), arC: [124.27 (C),
124.46 (CH), 127.10 (2CH), 128.21 (CH), 129.42 (CH), 135.92 (2CH),
137.21 (C), 148.69 (CH), 151.59 (CH)], 145.03 (triazole C-3), 156.08
(triazole C-5). EI MS m/z (%): 253.08 ([Mþ1]þ, 40), 271.22
([MþH2O]þ, 34), 275.16 ([MþNa]þ, 62), 293.18 ([Mþ2þK]þ, 64),
185.96 (17), 163.93 (22), 161.93 (41), 148.86 (35), 137.91 (100),
120.89 (88), 107.88 (32), 107.01 (54). Elemental analysis, for
C14H12N4O2, calcd. (%): 66.65 C, 4.79 H, 22.21 N; Found (%): 66.51
C, 5.09 H, 22.05 N.
4-Benzyl-5-(pyridin-3-yl)-4H-1,2,4-triazole-3-thiol (5)
Yield: 68%, m.p. 173–174°C. IR (KBr, n, cmꢀ1): 2725 (SH), 1548
(C N). 1H NMR (DMSO-d6, d ppm): 5.37 (s, 2H, CH2), 7.00 (s, 2H,
–
–
4-Phenyl-5-(pyridin-3-yl)-4H-1,2,4-triazole-3-thiol (7)
arH), 7.20 (d, 3H, arH, J ¼ 8.2 Hz), 7.46 (brs, 1H, arH), 7.93 (s, 1H,
arH), 8.66 (s, 2H, arH), 14.30 (brs, 1H, SH). 13C NMR (DMSO-d6,
d ppm): 47.37 (CH2), arC: [123.16 (C), 124.46 (2CH), 127.21 (CH),
128.32 (CH), 129.34 (CH), 136.18 (C), 136.70 (2CH), 149.25 (CH),
152.16 (CH)], 149.87 (triazole C-3), 169.08 (triazole C-5). EI MS m/z
(%): 269.22 ([Mþ1]þ, 88), 270.22 ([Mþ2]þ, 18), 179.03 (64), 177.96
(100), 150.03 (38), 132.93 (56), 120.03 (24). Elemental analysis,
for C14H12N4S, calcd. (%): 62.66 C, 4.51 H, 20.88 N; Found (%): 62.57
C, 4.53 H, 20.87 N.
Yield: 74%, m.p. 269–270°C. IR (KBr, n, cmꢀ1): 2763 (SH), 1578
(C N). 1H NMR (DMSO-d6, d ppm): 7.34–7.48 (m, 6H, arH), 7.66
–
–
(d, 1H, arH, J ¼ 7.8 Hz), 8.50–8.59 (m, 2H, arH), 14.30 (s, 1H, SH).
13C NMR (DMSO-d6, d ppm): arC: [122.87 (C), 124.47 (CH), 129.23
(2CH), 130.29 (2CH), 130.57 (CH), 134.47 (C), 136.94 (CH), 148.96
(CH), 151.59 (CH)], 149.31 (triazole C-3), 169.16 (triazole C-5). EI MS
m/z (%): 255.13 ([Mþ1]þ, 100), 256.14 ([Mþ2]þ, 18), 175.94 (18),
148.90 (18). Elemental analysis, for C13H10N4S, calcd. (%): 61.40 C,
3.96 H, 22.03 N; Found (%): 61.14 C, 4.28 H, 21.67 N.
4-Benzyl-5-pyridin-3-yl-2,4-dihydro-3H-1,2,4-triazol-3-one (6) General method for the synthesis of compounds 8–11
Yield: 66%, m.p. 165–167°C. IR (KBr, n, cmꢀ1): 3159 (NH), 1698
To a solution of corresponding compound 5, 6, or 7 (10 mmol) in
absolute ethanol, sodium (10 mmol) was added and the mixture
1
–
–
(C O), 1574 (C N). H NMR (DMSO-d , d ppm): 4.97 (s, 2H, CH ),
–
–
6
2
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Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
Hybrid Molecules Containing Different Heterocyclic Moieties
9
was stirred at room temperature for 2 h. Then, suitable alkyl
halide (20 mmol) was added into it and refluxed for 10 h (the
progress of the reaction was monitored by TLC). After evaporating
the solvent under reduced pressure, a solid appeared. This was
recrystallized from ethanol (for 8, 9, 11) or ethyl acetate/diethyl
ether (1:3; for 10) to afford the desired compound.
General method for the synthesis of compounds 12–24
To a solution of corresponding compound 5, 6, or 7 (10 mmol) in
tetrahydrofuran, suitable primary or secondary amine (10 mmol)
was added and the mixture was stirred at room temperature in
the presence of formaldehyde (37%, 3.72 mL, 5 mmol) for 2 h (the
progress of the reaction was monitored by TLC). After evaporating
the solvent under reduced pressure, a solid appeared. The crude
product was recrystallized from ethanol/water (1:3; for 12, 13, 19,
20–22), ethyl acetate (for 14), benzene/petroleum ether (1:2; for
15–18, 24), or ether (for 23) to yield the target compounds.
3-[4-Benzyl-5-(methylsulfanyl)-4H-1,2,4-triazol-3-yl]-
pyridine (8)
1
Yield: 57%, m.p. 90–92°C. IR (KBr, n, cmꢀ1): 1469 (C N). H NMR
–
–
(DMSO-d6, d ppm): 2.63 (s, 3H, CH3), 5.26 (s, 2H, CH2), 6.95 (s, 2H,
arH), 7.26 (s, 3H, arH), 7.50 (brs, 1H, arH), 7.96 (d, 1H, arH,
J ¼ 6.6 Hz), 8.69 (d, 2H, arH, J ¼ 8.2 Hz). 13C NMR (DMSO-d6, d ppm):
15.80 (CH3), 48.07 (CH2), arC: [123.98 (C), 124.62 (CH), 126.82
(2CH), 128.62 (CH), 129.63 (2CH), 135.84 (C), 136.55 (CH), 149.19
(CH), 151.63 (CH)], 153.71 (triazole C-3), 170.01 (triazole C-5). EI MS
m/z (%): 305.26 ([MþNa]þ, 100), 306.26 ([Mþ1þNa]þ, 19). Elemental
analysis, for C15H14N4S, calcd. (%): 63.80 C, 5.00 H, 19.84 N; Found
(%): 63.52 C, 5.28 H, 19.67 N.
4-Benzyl-2-(morpholin-4-ylmethyl)-5-(pyridin-3-yl)-2,4-
dihydro-3H-1,2,4-triazole-3-thione (12)
Yield: 80%, m.p. 130–132°C. IR (KBr, n, cmꢀ1): 1578 (C N), 1159
–
–
1
–
–
(C S), 1112 (C–O). H NMR (DMSO-d , d ppm): 2.72 (s, 4H, 2NCH ),
6
2
3.55 (s, 4H, 2OCH2), 5.16 (s, 2H, benzyl-CH2), 5.54 (s, 2H, CH2),
7.00 (s, 2H, arH), 7.21 (s, 3H, arH), 7.46 (d, 1H, arH, J ¼ 7.4 Hz), 7.95
(d, 1H, arH, J ¼ 7.8 Hz), 8.66 (s, 2H, arH). 13C NMR (DMSO-d6,
d ppm): 48.46 (benzyl-CH2), 51.02 (2NCH2), 66.78 (2OCH2), 69.99
(CH2), arC: [104.99 (CH), 122.75 (C), 124.50 (CH), 127.14 (2CH),
128.37 (CH), 129.39 (CH), 136.06 (C), 139.91 (CH), 149.38 (CH),
152.39 (CH)], 148.71 (triazole C-3), 170.09 (triazole C-5). EI MS
m/z (%): 368.27 ([Mþ1]þ, 39), 369.27 ([Mþ2]þ, 14), 383.35
([Mꢀ2þH2O]þ, 21), 385.23 ([MþH2O]þ, 16), 327.23 (69), 323.22
(100), 301.20 (66), 269.10 (58), 196.83 (36), 132.82 (31), 118.37 (31),
113.86 (32). Elemental analysis, for C19H21N5OS, calcd. (%): 62.10
C, 5.76 H, 19.06 N; Found (%): 62.47 C, 5.79 H, 19.24 N.
3-[5-(Methylsulfanyl)-4-phenyl-4H-1,2,4-triazol-3-yl]-
pyridine (9)
1
Yield: 53%. m.p. 165–167°C. IR (KBr, n, cmꢀ1): 1498 (C N). H NMR
–
–
(DMSO-d6, d ppm): 2.63 (s, 3H, CH3), 7.52 (d, 6H, arH, J ¼ 14.0 Hz),
7.72 (d, 1H, arH, J ¼ 6.2 Hz), 8.55 (s, 2H, arH). 13C NMR (DMSO-d6,
d ppm): 15.09 (CH3), arC: [123.71 (C), 124.28 (CH), 128.34 (2CH),
130.80 (2CH), 130.99 (CH), 134.11 (C), 136.07 (CH), 148.96 (CH),
151.15 (CH)], 152.94 (triazole C-3), 154.18 (triazole C-5). EI MS m/z
(%): 269.15 ([Mþ1]þ, 100), 291.17 ([MþNa]þ, 62), 270.10 ([Mþ2]þ,
17). Elemental analysis, for C14H12N4S, calcd. (%): 62.66 C, 4.51 H,
20.88 N; Found (%): 62.55 C, 4.65 H, 20.53 N.
4-Benzyl-5-(pyridin-3-yl)-2-(thiomorpholin-4-ylmethyl)-2,4-
dihydro-3H-1,2,4-triazole-3-thione (13)
Yield: 83%, m.p. 127–129°C. IR (KBr, n, cmꢀ1): 1595 (C N), 1137
–
–
1
–
–
(C S). H NMR (DMSO-d , d ppm): 2.62 (s, 4H, N-2CH ), 3.03 (s, 4H,
6
2
3-[5-(Hexylsulfanyl)-4-phenyl-4H-1,2,4-triazol-3-yl]-
S-2CH2), 5.19 (s, 2H, benzyl-CH2), 5.43 (s, 2H, CH2), 7.01 (s, 2H, arH),
7.23 (s, 3H, arH), 7.51 (d, 1H, arH, J ¼ 4.0 Hz), 7.97 (d, 1H, arH,
J ¼ 8.0 Hz), 8.70 (s, 2H, arH). 13C NMR (DMSO-d6, d ppm): 27.84
(N-2CH2), 48.45 (benzyl-CH2), 53.06 (S-2CH2), 71.39 (CH2),
arC: [122.75 (C), 124.53 (CH), 127.14 (2CH), 128.35 (CH), 129.45
(2CH), 136.06 (C), 136.88 (CH), 149.35 (CH), 152.41 (CH)], 148.73
(triazole C-3), 169.87 (triazole C-5). EI MS m/z (%): 413.39 (46),
383.80 ([M]þ, 30), 269.10 (55), 180.87 (34), 178.94 (100), 162.85
(30), 119.93 (34), 104.91 (44). Elemental analysis, for C19H21N5S2,
calcd. (%): 59.50 C, 5.52 H, 18.26 N; Found (%): 59.55 C, 5.42
H, 18.58 N.
pyridine (10)
1
Yield: 61%, m.p. 65–67°C. IR (KBr, n, cmꢀ1): 1592 (C N). H NMR
–
–
(DMSO-d6, d ppm): 1.25 (brs, 9H, 3CH2 þ CH3), 1.68 (brs, 4H, 2CH2),
7.57 (brs, 5H, arH), 8.18 (brs, 2H, arH), 8.56 (brs, 1H, arH), 9.21
(d, 1H, arH, J ¼ 21.0 Hz). 13C NMR (DMSO-d6, d ppm): 14.60 (CH3),
25.58 (CH2), 28.32 (CH2), 29.60 (CH2), 31.37 (CH2), 32.74 (CH2), arC:
[127.63 (C), 128.33 (CH), 128.45 (CH), 128.68 (CH), 130.72 (CH),
130.95 (CH), 131.34 (CH), 133.25 (C), 136.06 (CH), 143.67 (CH),
148.98 (CH)], 150.02 (triazole C-3), 154.73 (triazole C-5). EI MS m/z
(%): 339.30 ([Mþ1]þ, 100), 340.30 ([Mþ2]þ, 24), 255.15 (48).
Elemental analysis, for C19H22N4S, calcd. (%): 67.42 C, 6.55 H,
16.55 N; Found (%): 67.02 C, 6.65 H, 16.20 N.
4-Benzyl-2-{[4-(4-fluorophenyl)piperazine-1-yl]methyl}-5-
(pyridine-3-yl)-2,4-dihydro-3H-1,2,4-triazole-3-thione (14)
Yield: 85%, m.p. 102–103°C. IR (KBr, n, cmꢀ1): 1567 (C N),
3-[5-(Benzylsulfanyl)-4-phenyl-4H-1,2,4-triazol-3-yl]-
–
–
1
pyridine (11)
–
1179 (C S). H NMR (DMSO-d , d ppm): 2.91 (s, 4H, N-2CH ), 3.07
–
6
2
1
Yield: 53%, m.p. 165–167°C. IR (KBr, n, cmꢀ1): 1496 (C N). H NMR
–
–
(s, 4H, N-2CH2), 5.26 (s, 2H, benzyl-CH2), 5.43 (s, 2H, CH2), 7.00
(d, 6H, arH, J ¼ 8.0 Hz), 7.22 (s, 3H, arH), 7.49 (brs, 1H, arH), 7.96
(d, 1H, arH, J ¼ 6.0 Hz), 8.70 (s, 2H, arH). 13C NMR (DMSO-d6,
d ppm): 49.77 (benzyl-CH2), 50.53 (N-2CH2), 56.75 (N-2CH2),
69.74 (CH2), arC: [115.73 (CH), 116.16 (CH), 118.02 (CH), 118.17
(CH), 122.73 (C), 124.53 (CH), 127.10 (2CH), 128.39 (CH), 129.41
(2CH), 136.01 (C), 136.91 (CH), 148.69 (C), 149.32 (CH), 152.39
(CH), 154.76 (C)], 159.10 (triazole C-3), 170.01 (triazole C-5).
EI MS m/z (%): 461.33 ([M þ 1]þ, 11), 235.00 (100), 429.42 (14),
414.40 (26), 413.40 (16). Elemental analysis, for C25H25FN6S,
calcd. (%): 65.19 C, 5.47 H, 18.25 N; Found (%): 65.05 C, 5.37 H,
18.10 N.
(DMSO-d6, d ppm): 5.89 (s, 2H, CH2), 7.24–7.50 (m, 12H, arH), 8.09–
8.17 (m, 1H, arH), 8.31 (d, 1H, arH, J ¼ 7.4 Hz). 13C NMR (DMSO-d6,
d ppm): 38.78 (benzyl-CH2), arC: [128.85 (CH), 129.90 (C), 130.25
(CH), 130.37 (CH), 130.48 (CH), 131.01 (CH), 131.20 (CH), 131.39
(CH), 131.58 (CH), 131.88 (CH), 132.17 (CH), 132.40 (CH), 132.97
(CH), 133.16 (CH), 133.63 (CH), 134.97 (C), 136.37 (C)], 139.55
(triazole C-3), 152.17 (triazole C-5). EI MS m/z (%): 345.31 ([Mþ1]þ,
100), 344.25 ([M]þ, 72), 367.27 ([MþNa]þ, 20), 374.40 (22), 373.34
(63), 346.31 (28), 343.37 (22), 276.98 (22). Elemental analysis, for
C20H16N4S, calcd. (%): 69.74 C, 4.68 H, 16.27 N; Found (%): 69.50 C,
4.38 H, 16.07 N.
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S. Ceylan et al.
Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
4-Benzyl-2-({[3-(1H-imidazol-1-yl)propyl]amino}methyl)-5-
(CH), 151.84 (CH), 154.42 (CH)], 155.26 (triazole C-3), 159.11
(triazole C-5). EI MS m/z (%): 445.36 ([Mþ1]þ, 24), 467.38 ([MþNa]þ,
14), 414.45 (26), 413.45 (100). Elemental analysis, for C25H25FN6O,
calcd. (%): 67.55 C, 5.67 H, 18.91 N; Found (%): 62.47 C, 5.82 H,
18.53 N.
(pyridin-3-yl)-2,4-dihydro-3H-1,2,4-triazole-3-thione (15)
Yield: % 71, m.p.129–131°C. IR (KBr, n, cmꢀ1): 3107 (NH), 1576
(C N) 1133 (C S). 1H NMR (DMSO-d6, d ppm): 3.23–3.55 (m, 2H,
–
–
–
–
CH2 þ H2O), 3.95 (brs, 4H, 2CH2), 5.40 (brs, 2H, CH2), 5.54 (brs, 2H,
CH2), 7.05 (d, 2H, arH, J ¼ 11 Hz), 7.14–7.34 (m, 4H, arH), 7.43–7.62
(m, 3H, arH), 7.89 (d, 1H, arH, J ¼ 8.2 Hz), 8.63–8.69 (m, 2H, arH),
10.57 (s, 1H, NH). 13C NMR (DMSO-d6, d ppm): 28.84 (CH2), 48.49
(benzyl-CH2), 56.74 (CH2), 67.28 (CH2), 74.07 (CH2), arC: [119.80
(CH), 122.62 (C), 124.48 (CH), 127.14 (2CH), 128.37 (CH), 128.91
(CH), 129.37 (2CH), 135.92 (C), 136.76 (CH), 149.29 (CH), 152.19
(CH), 152.41 (CH)], 148.94 (triazole C-3), 169.13 (triazole C-5). EI MS
m/z (%): 434.41 (100), 435.41 (26), 418.33 (46), 412.45 (81), 410.44
(18). Elemental analysis, for C21H23N7S, calcd. (%): 62.20 C, 5.72 H,
24.18 N; Found (%): 65.05 C, 5.37 H, 24.48 N.
2-(Morpholinomethyl)-4-phenyl-5-(pyridin-3-yl)-2H-1,2,4-
triazole-3(4H)-thione (19)
Yield: 87%, m.p. 209–210°C. IR (KBr, n, cmꢀ1): 1576 (C N), 1154
–
–
(C S), 1114 (C–O). 1H NMR (DMSO-d6, d ppm): 2.81 (brs, 4H,
–
–
N–2CH2), 3.60 (brs, 4H, O–2CH2), 5.18 (brs, 2H, CH2), 7.49 (brs, 6H,
arH), 7.70 (d, 1H, arH, J ¼ 8.2 Hz), 8.55 (d, 2H, arH, J ¼ 14.0 Hz).
13C NMR (DMSO-d6, d ppm): 50.94 (N–2CH2), 66.82 (O–2CH2), 69.83
(CH2), arC: [122.62 (C), 124.20 (CH), 129.49 (2CH), 130.16 (2CH),
130.45 (CH), 135.32 (C), 136.79 (CH), 149.50 (CH), 151.89 (CH)],
147.82 (triazole C-3), 170.42 (triazole C-5). EI MS m/z (%): 354.26
([Mþ1]þ, 66), 355.45 ([Mþ2]þ, 20), 377.41 ([Mþ1þNa]þ, 35), 385.54
(43), 383.54 (100), 383.41 (76), 379.41 (36), 348.50 (30), 338.49 (46),
336.42 (69), 334.36 (51), 332.36 (42), 255.21 (64), 255.08 (85),
253.14 (51), 252.96 (40). Elemental analysis, for C18H19N5OS,
calcd. (%): 61.17 C, 5.42 H, 19.81 N; Found (%): 61.28 C, 5.52 H,
19.92 N.
4-Benzyl-2-(morpholin-4-ylmethyl)-5-(pyridin-3-yl)-2,4-
dihydro-3H-1,2,4-triazol-3-one (16)
Yield: 84%, m.p.110–112°C. IR (KBr, n, cmꢀ1): 1705 (C O), 1572
–
–
1
–
–
(C N), 1114 (C–O). H NMR (DMSO-d , d ppm): 2.62 (s, 4H, 2NCH ),
6
2
3.41 (s, 4H, 2OCH2), 4.69 (s, 2H, benzyl-CH2), 5.01 (s, 2H, CH2), 7.05
(d, 1H, arH, J ¼ 7.0 Hz), 7.26 (d, 4H, arH, J ¼ 7.0 Hz), 7.44–7.50
(m, 1H, arH), 7.94 (d, 1H, arH, J ¼ 7.2 Hz), 8.64–8.70 (m, 2H, arH).
13C NMR (DMSO-d6, d ppm): 45.40 (benzyl-CH2), 50.65 (N-2CH2),
66.75 (O–2CH2), 66.89 (CH2), arC: [123.79 (C), 124.51 (CH), 127.06
(2CH), 128.31 (CH), 129.48 (2CH), 136.09 (CH), 137.00 (C), 148.78
(CH), 151.83 (CH)], 143.75 (triazole C-3), 155.26 (triazole C-5). EI MS
m/z (%): 374.31 ([MþNa]þ, 15), 249.15 (47). Elemental analysis, for
C19H21N5O2, calcd. (%): 64.94 C, 6.02 H, 19.93 N; Found (%): 64.57
C, 6.25 H, 19.61 N.
4-Phenyl-5-(pyridin-3-yl)-2-(thiomorpholin-4-ylmethyl)-2,4-
dihydro-3H-1,2,4-triazole-3-thione (20)
Yield: 88%, m.p. 175–176°C. IR (KBr, n, cmꢀ1): 1575 (C N), 1144
–
–
1
–
–
(C S). H NMR (DMSO-d , d ppm): 2.64 (brs, 4H, N–2CH ), 3.10 (brs,
6
2
4H, S–2CH2), 5.19 (s, 2H, CH2), 7.44–7.50 (m, 6H, arH), 7.70 (d, 1H,
arH, J ¼ 8.2 Hz), 8.52 (s, 1H, arH), 8.60 (d, 1H, arH, J ¼ 4.6 Hz).
13C NMR (DMSO-d6, d ppm): 27.88 (N–2CH2), 52.92 (S–2CH2), 71.21
(CH2), arC: [122.62 (C), 124.21 (CH), 129.50 (2CH), 130.16 (2CH),
130.46 (CH), 135.31 (C), 136.75 (CH), 149.48 (CH), 151.91 (CH)],
147.82 (triazole C-3), 170.22 (triazole C-5). EI MS m/z (%): 370.34
([Mþ1]þ, 28), 345.18 (20), 337.17 (26), 301.26 (21), 255.15 (100).
Elemental analysis, for C18H19N5S2, calcd. (%): 58.51 C, 5.18 H,
18.95 N; Found (%): 58.18 C, 5.41 H, 18.55 N.
4-Benzyl-5-(pyridin-3-yl)-2-(thiomorpholin-4-ylmethyl)-2,4-
dihydro-3H-1,2,4-triazol-3-one (17)
Yield: 82%, m.p.103–104°C. IR (KBr, n, cmꢀ1): 1699 (C O), 1574
–
–
1
–
–
(C N). H NMR (DMSO-d , d ppm): 2.61 (s, 4H, N–2CH ), 2.90 (s, 4H,
6
2
S–2CH2), 4.71 (s, 2H, benzyl-CH2), 5.01 (s, 2H, CH2), 7.05 (d, 2H,
arH, J ¼ 5.8 Hz), 7.26 (d, 3H, arH, J ¼ 7.0 Hz), 7.48 (brs, 1H, arH),
7.95 (d, 1H, arH, J ¼ 7.6 Hz), 8.70 (s, 2H, arH). 13C NMR (DMSO-d6,
d ppm): 27.87 (N–2CH2), 41.43 (benzyl-CH2), 52.72 (S–2CH2), 68.31
(CH2), arC: [123.79 (C), 124.52 (CH), 127.05 (2CH), 128.31 (CH),
129.02 (CH), 129.48 (CH), 136.06 (CH), 137.03 (C), 148.78 (CH),
151.84 (CH)], 143.70 (triazole C-3), 155.22 (triazole C-5). EI MS m/z
(%): 368.27 ([Mþ1]þ, 62), 369.34 ([Mþ2]þ, 19), 391.30 ([Mþ1þNa]þ,
24), 390.30 (100), 352.32 (20), 326.41 (25), 275.11 (37), 253.14 (31),
249.14 (76), 246.20 (21), 210.16 (54), 210.03 (51), 209.09 (46),
201.02 (26), 189.07 (19), 170.80 (26). Elemental analysis, for
C19H21N5OS, calcd. (%): 62.10 C, 5.76 H, 19.06 N; Found (%): 61.81
C, 5.55 H, 18.87 N.
4-Phenyl-2-{[4-(4-fluorophenyl)piperazine-1-yl]methyl}-5-
(pyridine-3-yl)-2,4-dihydro-3H-1,2,4-triazole-3-thione (21)
Yield: 89%, m.p. 171–172°C. IR (KBr, n, cmꢀ1): 1577 (C N), 1173
–
–
(C S). 1H NMR (DMSO-d6, d ppm): 2.99 (brs, 4H, N–2CH2), 3.09
–
–
(s, 4H, N–2CH2), 5.27 (s, 2H, CH2), 6.95–7.08 (m, 4H, arH), 7.49 (brs,
6H, arH), 7.70 (d, 1H, arH, J ¼ 8.2 Hz), 8.57 (d, 2H, arH, J ¼ 14.0 Hz).
13C NMR (DMSO-d6, d ppm): 49.85 (2NCH2), 50.84 (2NCH2), 69.61
(CH2), arC: [115.74 (CH), 116.17 (CH), 118.02 (CH), 118.17 (CH),
122.64 (C), 124.18 (CH), 129.50 (2CH), 130.16 (2CH), 130.43 (CH),
135.35 (C), 136.77 (CH), 147.80 (C), 148.72 (C), 149.52 (CH), 151.89
(CH)], 159.13 (triazole C-3), 170.39 (triazole C-5). EI MS m/z (%):
446.54 ([M]þ, 36), 414.46 (17), 413.46 (51), 227.61 (66), 137.96 (100),
181.07 (68). Elemental analysis, for C24H23FN6S, calcd. (%): 64.55 C,
5.19 H, 18.82 N; Found (%): 64.31 C, 5.11 H, 18.79 N.
4-Benzyl-2-{[4-(4-fluorophenyl)piperazine-1-yl]methyl}-5-
(pyridine-3-yl)-2,4-dihydro-3H-1,2,4-triazole-3-one (18)
Yield: 89%, m.p. 120–121°C. IR (KBr, n, cmꢀ1): 1699 (C O), 1511
–
–
1
4-Phenyl-2-({[3-(1H-imidazol-1-yl)propyl]amino}methyl)-5-
–
(C N). H NMR (DMSO-d , d ppm): 2.79 (s, 4H, N–2CH ), 3.07 (s, 4H,
–
6
2
(pyridin-3-yl)-2,4-dihydro-3H-1,2,4-triazole-3-thione (22)
N–2CH2), 4.78 (s, 2H, benzyl-CH2), 5.02 (s, 2H, CH2), 6.93–7.07
(m, 6H, arH), 7.26 (d, 3H, arH, J ¼ 7.0 Hz), 7.47 (t, 1H, arH,
J ¼ 4.8 Hz), 7.94 (d, 1H, arH, J ¼ 8.6 Hz), 8.65–8.70 (m, 2H, arH). 13C
NMR (DMSO-d6, d ppm): 45.42 (benzyl-CH2), 49.68 (N–2CH2), 50.24
(N–2CH2), 66.64 (CH2), arC: [115.73 (CH), 116.16 (CH), 117.98 (CH),
118.12 (CH), 123.81 (C), 124.51 (CH), 127.08 (2CH), 128.32 (CH),
129.49 (CH), 136.09 (CH), 137.03 (C), 143.76 (C), 148.61 (C), 148.81
Yield: 84%, m.p. 203–204°C. IR (KBr, n, cmꢀ1): 3049 (NH), 1578
1
–
–
(C N), 1140 (C S). H NMR (DMSO-d , d ppm): 2.94 (brs, 2H, CH ),
–
–
6
2
3.39 (brs, 2H, CH2 þ H2O), 4.04 (brs, 2H, CH2), 5.53 (s, 2H, CH2), 6.82
(s, 1H, arH), 7.15 (s, 1H, arH), 7.42–7.65 (m, 9H, arH), 8.48 (s, 1H,
arH), 8.59 (d, 1H, NH, J ¼ 3.8 Hz). 13C NMR (DMSO-d6, d ppm): 28.19
(CH2), 43.78 (CH2), 46.13 (CH2), 66.91 (CH2), arC: [119.90 (CH),
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Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
Hybrid Molecules Containing Different Heterocyclic Moieties
11
122.55 (C), 124.20 (2CH), 129.39 (2CH), 130.19 (2CH), 130.52 (2CH),
135.16 (C), 136.67 (CH), 149.45 (CH), 151.92 (CH)], 148.03 (triazole
C-3), 169.46 (triazole C-5). EI MS m/z (%): 412.51 ([Mꢀ2þNa]þ, 23),
434.53 (30), 405.38 (25), 434.38 (100). Elemental analysis, for
443.23 ([Mꢀ1þNa]þ, 10), 459.25 ([Mꢀ1þK]þ, 33), 393.05 (100),
435.22 (27), 395.05 (70). Elemental analysis, for C22H17ClN4OS,
calcd. (%): 62.78 C, 4.07 H, 13.31 N; Found (%): 62.45 C, 4.36 H,
13.61 N.
C20H21N7S, calcd. (%): 61.36 C, 5.41 H, 25.04 N; Found (%): 61.51 C,
5.11 H, 25.36 N.
N0-[3-Benzyl-4-(4-chlorophenyl)-1,3-oxazol-2(3H)-ylidene]
pyridine-3-carbohydrazide (26)
Yield: 50%, m.p. 98–100°C. IR (KBr, n, cmꢀ1): 3263 (NH), 1641
2-[(Pentylamino)methyl]-4-phenyl-5-(pyridin-3-yl)-2,4-
(C O), 1531 (C N). 1H NMR (DMSO-d6, d ppm): 4.24 (d, 2H, CH2,
–
–
–
–
dihydro-3H-1,2,4-triazole-3-thione (23)
Yield: 81%, m.p. 229–231°C. IR (KBr, n, cmꢀ1): 3050 (NH), 1576
J ¼ 5.4 Hz), 7.27–7.61 (m, 9H, arH þ oxazole-CH), 7.95–8.21 (m, 3H,
1
–
–
(C N), 1192 (C S). H NMR (DMSO-d , d ppm): 0.85 (brs, 2H, CH ),
arH), 8.73 (s, 1H, arH), 9.05 (s, 1H, arH), 9.87 (s, 1H, NH). 13C NMR
–
–
6
2
1.26 (brs, 3H, CH3), 1.52 (brs, 2H, CH2), 2.75 (d, 2H, CH2, J ¼ 7.0 Hz),
4.17 (brs, 2H, CH2 þ H2O), 5.58 (s, 2H, CH2), 7.47 (d, 6H, arH,
J ¼ 11.8 Hz), 7.79 (d, 1H, arH, J ¼ 7.0 Hz), 8.56 (s, 1H, arH), 8.65
(d, 1H, arH, J ¼ 4.0 Hz), 10.33 (s, 1H, NH). 13C NMR (DMSO-d6,
d ppm): 14.40 (CH2), 14.47 (CH3), 22.33 (CH2), 27.32 (CH2), 28.60
(CH2), 71.64 (CH2), arC: [123.11 (C), 124.87 (CH), 129.42 (CH),
130.26 (CH), 130.56 (CH), 134.97 (C), 138.21 (CH), 147.79 (CH),
148.27 (CH), 150.07 (CH), 150.71 (CH)], 150.78 (triazole C-3), 169.16
(triazole C-5). EI MS m/z (%): 367.51 ([M]þ, 44), 301.22 (91), 255.14
(24), 188.93 (48), 180.97 (34), 170.94 (30), 148.93 (100), 118.84 (76),
116.89 (36). Elemental analysis, for C19H23N5S, calcd. (%): 64.56 C,
6.56 H, 19.81 N; Found (%): 64.40 C, 6.22 H, 19.61 N.
(DMSO-d6, d ppm): 67.10 (CH2), 124.17 (oxazole C-5), arC:
[127.20 (CH), 127.59 (CH), 127.98 (CH), 128.80 (CH), 129.13 (C),
129.64 (2CH), 129.77 (CH), 130.42 (CH), 130.80 (CH), 131.57 (CH),
136.06 (CH), 139.21 (C), 141.31 (C), 149.37 (CH), 152.93 (CH),
165.77 (C)], 159.08 (oxazole C-2), 167.17 (þoxazole C-4), 168.57
–
(C O). EI MS m/z (%): 445.30 ([Mþ2þK] , 19), 419.45 (36),
–
414.51 (25), 413.45 (100). Elemental analysis, for C22H17ClN4O2,
calcd. (%): 65.27 C, 4.23 H, 13.84 N; Found (%): 65.35 C, 4.60 H,
13.57 N.
N0-[5-(4-chlorophenyl)-3-phenyl-1,3-thiazole-2(3H)-
ylidene]pyridine-3-carbo-hydrazide (27)
Yield: 60%, m.p. 226–227°C. IR (KBr, n, cmꢀ1): 3268 (NH), 1630
1
2-{[(Furan-2-ylmethyl)amino]methyl}-4-phenyl-5-(pyridin-
–
–
(C O), 1555 (C N). H NMR (DMSO-d , d ppm): 6.98–7.05 (m, 1H,
–
–
6
3-yl)-2,4-dihydro-3H-1,2,4-triazole-3-thione (24)
thiazole-CH), 7.33–7.40 (m, 3H, arH), 7.62 (d, 4H, arH, J ¼ 7.0 Hz),
8.24 (d, 2H, arH, J ¼ 4.2 Hz), 8.75 (s, 2H, arH), 9.06 (s, 2H, arH),
10.80 (s, 1H, NH). 13C NMR (DMSO-d6, d ppm): 143.56 (thiazole-CH),
arC: [117.83 (2CH), 121.11 (C), 122.73 (2CH), 125.02 (2CH), 129.83
(2CH), 133.78 (2CH), 139.18 (2C), 146.94 (2CH), 150.56 (C), 152.19
–
Yield: 79%, m.p. 167–168°C. IR (KBr, n, cmꢀ1): 3139 (NH), 1785
(C N), 1146 (C S), 1127 (C–O). 1H NMR (DMSO-d6, d ppm): 4.41
–
–
–
–
(brs, 2H, CH2), 5.64 (brs, 2H, CH2), 6.42 (brs, 1H, NH), 7.50 (brs, 9H,
arH), 8.44–8.60 (m, 3H, arH). 13C NMR (DMSO-d6, d ppm): 67.55
(CH2), 71.59 (CH2), arC: [122.58 (C), 124.31 (2CH), 129.21 (CH),
129.40 (2CH), 130.27 (2CH), 130.56 (CH), 135.05 (C), 136.65 (CH),
136.83 (CH), 148.12 (C), 149.39 (CH), 151.92 (CH)], 152.62 (triazole
C-3), 169.14 (triazole C-5). EI MS m/z (%): 363.44 ([M]þ, 56), 414.51
(28), 413.51 (100), 376.34 (22), 258.34 (56), 255.15 (26), 152.91 (13).
Elemental analysis, for C19H17N5OS, calcd. (%): 62.79 C, 4.71 H,
19.27 N; Found (%): 62.45 C, 4.50 H, 19.50 N.
(CH)], 156.56 (thiazole C-5), 160.91 (thiazole C-2), 172.33 (C O).
–
EI MS m/z (%): 413.45 (100), 414.45 (27). Elemental analysis, for
C21H15ClN4OS, calcd. (%): 61.99 C, 3.72 H, 13.77 N; Found (%): 61.60
C, 3.40 H, 13.42 N.
General method for the synthesis of compounds 28–30
The mixture of corresponding compound 2, 3, or 4 (10 mol) and
ethyl bromoacetate (10 mmol) in absolute ethanol was refluxed in
the presence of sodium acetate (50 mmol) for 8 h (the progress of
the reaction was monitored by TLC). After evaporating the solvent
under reduced pressure, a solid appeared. The crude product was
washed with water and recrystallized from ethyl acetate/
petroleum ether (1:3; for 28), dimethyl sulfoxide/water (1:3; for
29), or ethanol (for 30) a suitable solvent to afford the desired
compound.
General method for the synthesis of compounds 25–27
4-Chlorophenacylbromide (10 mmol) and sodium acetate
(50 mmol) were added to the solution of the corresponding
compound 2, 3, or 4 (10 mmol) in ethanol and the reaction
mixture was allowed to reflux for 8 h (the progress of the
reaction was monitored by TLC). Then, the mixture was cooled to
room temperature and left overnight in the cold. The formed
solid was filtered, washed with water, and recrystallized from
ethyl acetate (for 25) or ethanol (for 26, 27) to afford the pure
compound.
N0-[3-Benzyl-5-oxo-1,3-thiazolidin-2-ylidene]pyridine-3-
carbohydrazide (28)
Yield: 51%, m.p. 160–162°C. IR (KBr, n, cmꢀ1): 3202 (NH), 1722
N0-[3-Benzyl-5-(4-chlorophenyl)-1,3-thiazol-2(3H)-ylidene]-
1
–
–
(2C O), 1602 (C N). H NMR (DMSO-d , d ppm): 3.70 (s, 2H, CH ),
–
–
6
2
pyridine-3-carbohydrazide (25)
4.46 (s, 2H, benzyl-CH2), 6.85–7.06 (m, 6H, arH), 7.73 (s, 1H, arH),
8.26 (s, 1H, arH), 8.53 (s, 1H, arH), 10.67 (s, 1H, NH). 13C NMR
(DMSO-d6, d ppm): 46.75 (benzyl-CH2), 60.50 (thiazolidine C-4), arC:
[124.29 (CH), 124.95 (CH), 128.13 (CH), 128.26 (CH), 128.50 (CH),
129.12 (CH), 129.86 (C), 135.85 (CH), 136.55 (C), 149.05 (CH), 152.68
Yield: 57%, m.p. 185–187°C. IR (KBr, n, cmꢀ1): 3224 (NH), 1613
(C O), 1554 (C N). 1H NMR (DMSO-d6, d ppm): 4.45 (d, 2H, CH2,
–
–
–
–
J ¼ 5.8 Hz), 7.29–7.89 (m, 10H, arH þ thiazole-CH), 8.15 (d, 1H, arH,
J ¼ 7.8 Hz), 8.55 (s, 1H, arH), 8.69 (s, 1H, arH), 8.98 (s, 1H, arH),
10.28 (s, 1H, NH). 13C NMR (DMSO-d6, d ppm): 46.72 (CH2), 124.94
(thiazole-CH), arC: [121.33 (2C), 127.88 (CH), 128.11 (2CH), 129.07
(2CH), 130.27 (CH), 133.31 (2CH), 134.61 (CH), 139.32 (2C), 143.90
(CH)], 162.44 (thiazolidine C-2), 163.08 (thiazolidine C-5), 172.55
þ
–
(hydrazide C O). EI MS m/z (%): 327.10 ([Mþ1] , 42), 349.19
–
([MþNa]þ, 100), 350.13 ([Mþ1þNa]þ, 21), 229.12 (16). Elemental
analysis, for C16H14N4O2S, calcd. (%): 58.88 C, 4.32 H, 17.17 N;
Found (%): 58.60 C, 4.10 H, 17.42 N.
(CH), 146.59 (2CH), 151.78 (CH)], 156.56 (thiazole C-2), 160.91
þ
–
(thiazole C-5), 176.33 (C O). EI MS m/z (%): 439.23 ([MþH O] , 22),
–
2
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12
S. Ceylan et al.
Arch. Pharm. Chem. Life Sci. 2013, 346, 1–14
N0-[3-Benzyl-5-oxo-1,3-oxazolidin-2-ylidene]pyridine-3-
Urease inhibition assay [67]
Reaction mixtures comprising 25 mL of Jack bean urease, 55 mL of
buffer (100 mM urea, 0.01 M K2HPO4, 1 mM EDTA, and 0.01 M
LiCl2, pH 8.2), and 100 mM urea were incubated with 5 mL of
the test compounds at room temperature for 15 min in
microtiter plates. The production of ammonia was measured
by indophenol method and used to determine the urease
inhibitory activity. The phenol reagent (45 mL, 1% w/v phenol
and 0.005% w/v sodium nitroprusside) and alkali reagent (70 mL,
0.5% w/v sodium hydroxide and 0.1% NaOCl) were added to
each well and the increasing absorbance at 630 nm was
measured after 20 min, using a microplate reader (Molecular
Device, USA). The percentage inhibition was calculated from the
formula 100 ꢀ (ODtestwell/ODcontrol) ꢂ 100. Thiourea was used as
the standard inhibitor. In order to calculate IC50 values, different
concentrations of synthesized compounds and standard were
assayed under the same reaction conditions.
carbohydrazide (29)
Yield: 58%, m.p. 98–100°C. IR (KBr, n, cmꢀ1): 3211 (NH), 1683 and
1
–
–
1671 (2C O), 1595 (C N), 1116 (C–O). H NMR (DMSO-d , d ppm):
–
–
6
3.38 (s, 2H, CH2 þ H2O), 4.24 (d, 2H, benzyl-CH2, J ¼ 4.6 Hz),
7.27 (brs, 4H, arH), 7.53 (d, 1H, arH, J ¼ 4.8 Hz), 8.11 (s, 1H, arH),
8.23 (d, 1H, arH, J ¼ 7.4 Hz), 8.73 (s, 1H, arH), 9.05 (s, 1H,
arH), 10.39 (brs, 1H, NH). 13C NMR (DMSO-d6, d ppm): 43.35 (benzyl
CH2), 44.48 (oxazolidinone C-4), arC: [124.18 (CH), 127.21 (CH),
127.59 (2CH), 128.81 (2CH), 129.09 (2C), 136.07 (CH), 149.36 (CH),
152.90 (CH)], 141.28 (oxazolidinone C-2), 159.08 (oxazolidinone
þ
–
C-5), 165.82 (C O). EI MS m/z (%): 333.24 ([MþNa] , 10), 332.24
–
([Mꢀ1þNa]þ, 65), 294.19 (16), 293.18 (100), 246.07 (16),
165.88 (24), 137.89 (29). Elemental analysis, for C16H14N4O3,
calcd. (%): 61.93 C, 4.55 H, 18.06 N; Found (%): 61.60 C, 4.40 H,
17.51 N.
N0-[4-Oxo-3-phenyl-1,3-thiazolidine-2-ylidene]pyridine-3-
Antilipase activity assay
carbohydrazide (30)
The inhibitory effects of those compounds were evaluated against
porcine pancreatic lipase (PPL) (15 ng/mL). Lipase activity assay
was done according to Woods et al. [68]. Microtiter plates were
coated with purified tung oil TAGs. Compounds were mixed with
PPL 1:2 v/v and incubated for 30 min. The microtiter plates
containing purified tung oil, lipase solution, and assay buffer
(10 mM Tris–HCl buffer, pH 8.0, containing 150 mM NaCl, 6 mM
CaCl2, 1 mM EDTA, and 3 mg/mL b-cyclodextrin) were recorded
continuously for 40 min against the buffer alone by using
microplate reader (SpectraMax M5, Molecular Devices) at 272 nm.
The inhibitory activity of those compounds and orlistat, a positive
control against pancreatic lipase, was measured at concentra-
tions of 6.25, 2.08, and 1.04 mg/mL. Residual activities were
calculated by comparing with control without inhibitor (Tþ). The
assays were done in triplicate. The IC50 value was determined as
the concentration that gave 50% inhibition of maximal activity.
Yield: 67%, m.p. 228–230°C. IR (KBr, n, cmꢀ1): 3207 (NH), 1630
(C O), 1613 (C O), 1555 (C N). 1H NMR (DMSO-d6, d ppm): 3.39
–
–
–
–
–
–
(s, 2H, CH2 þ H2O), 6.98–7.05 (m, 1H, arH), 7.33–7.40 (m, 2H, arH),
7.62 (d, 3H, arH, J ¼ 7.4 Hz), 8.25 (d, 1H, arH, J ¼ 7.0 Hz), 8.74 (d, 1H,
arH, J ¼ 4.0 Hz), 9.06 (s, 1H, arH), 10.80 (s, 1H, NH). 13C NMR
(DMSO-d6, d ppm): 31.38 (thiazolidine C-5), arC: [117.84 (2CH),
121.10 (C), 122.74 (CH), 125.02 (CH), 129.83 (CH), 133.78 (CH),
139.17 (C), 146.96 (2CH), 152.21 (CH)], 156.56 (thiazolidine C-2),
–
160.91 (thiazolidine C-4), 207.29 (C O). EI MS m/z (%): 301.23 (100),
–
239.09 (62), 148.90 (58), 134.94 (23), 118.90 (29). Elemental
analysis, for C15H12N4O2S, calcd. (%): 57.68 C, 3.87 H, 17.94 N;
Found (%): 57.42 C, 4.11 H, 17.62 N.
Biological activity studies
Antimicrobial activity
The test microorganisms were obtained from the Hifzissihha
Institute of Refik Saydam (Ankara, Turkey) and were as follows:
Escherichia coli ATCC35218, Yersinia pseudotuberculosis ATCC911,
Pseudomonas aeruginosa ATCC43288, Enterococcus faecalis ATCC29212,
Staphylococcus aureus ATCC25923, Bacillus cereus 709 Roma, Mycobac-
terium smegmatis ATCC607, Candida albicans ATCC60193, and
Saccharomyces cerevisiae RSKK 251, Ar: A. oxydans (laboratory strain),
Ct: C. tropicalis, ATCC 13803, Pv: P. vulgaris ATCC 13315, Ac:
Acinetobacter sp. (laboratory strain), except S. marcescens (Sm),
Acinetobacter sp. (Ac) and K. oxitoka (Ko), which are laboratry
strains. All the newly synthesized compounds were weighed
and dissolved in hexane to prepare extract stock solution of
20.000 mg/mL.
The support provided by Karadeniz Technical University, BAP, Turkey
(Ref. No. 8623 and 8663), is gratefully acknowledged.
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