Synthesis and biological activity of 1,3,4-oxa(thia)diazole, 1,2,4-triazole-5-(thio)one and S-substituted derivatives of 3-((2-carboxyethyl)phenylamino)propanoic acid

Article abstract of DOI:10.1007/s11164-015-2290-0

Abstract: A series of novel derivatives of 3-((2-carboxyethyl)phenylamino)propanoic acid bearing two identical substituted 1,3,4-oxadiazole, 1,3,4-thiadiazole, or 1,2,4-triazole moieties were synthesized and their molecular structures were confirmed by IR, ¹H, ¹³C NMR spectroscopy, and elemental analysis data. The synthesized compounds were screened for their reducing power, free radical scavenging, plant growth regulating, and antimicrobial properties. 5,5′-((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-2,4-dihydro-3H-1,2,4-triazole-3-thione) showed excellent antioxidant activity, three times higher than that of the antibiotic control (cefazolin). 2,2′-((((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-4H-1,2,4-triazole-5,3-diyl))bis(sulfanediyl))diacetate exhibited notable bactericidal activity against Pseudomonas aeruginosa with MBC and MIC values 7.3?μg/ml, whereas 12 compounds displayed significant fungicidal activity against Candida albicans with MIC value 3.9?μg/ml. Graphical Abstract: Synthesis of compounds 2–27. R: 2, 5, 7, 10, 13, 15-27 C₆H₅–; 3, 8, 11 4-Cl-C₆H₄–; 4, 9, 12 CH₃–; 6, 14 2.6-(CH₃)₂-C₆H₃–; R′: 15 CH₃CH₂–; 16 (CH₃)₂CH–; 17 (CH₃)₂CHCH₂–; 18 CH₃CH₂CH₂CH₂–; 19 (CH₃)₂CHCH₂CH₂–; 20 CH₃CH₂CH₂CH₂CH₂–; 21 NH₂COCH₂–; 22 C₆H₅CH₂–; 23 C₆H₅COCH₂–; 24 4-ClC₆H₄COCH₂–; 25 4-NO₂C₆H₄COCH₂–; 26 C₂H₅OCOCH₂–.[Figure not available: see fulltext.]

Full text of DOI:10.1007/s11164-015-2290-0

Res Chem Intermed
DOI 10.1007/s11164-015-2290-0
Synthesis and biological activity of 1,3,4-
oxa(thia)diazole, 1,2,4-triazole-5-(thio)one
and S-substituted derivatives of 3-((2-
carboxyethyl)phenylamino)propanoic acid
1
I. Tumosiene I. Jonuskiene K. Kantminiene
1
2
•
•
•
_
ˇ
_
_
J. Siugzdaite V. Mickevicius Z. J. Beresnevicius
3
1
1
ˇ
•
•
_
ˇ
ˇ
ˇ
Received: 14 July 2015 / Accepted: 21 September 2015
Ó Springer Science+Business Media Dordrecht 2015
Abstract A series of novel derivatives of 3-((2-carboxyethyl)phenylamino)propanoic
acid bearing two identical substituted 1,3,4-oxadiazole, 1,3,4-thiadiazole, or 1,2,4-tria-
zole moieties were synthesized and their molecular structures were confirmed by IR, 1H,
13C NMR spectroscopy, and elemental analysis data. The synthesized compounds were
screened for their reducing power, free radical scavenging, plant growth regulating, and
antimicrobial properties. 5,50-((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-2,4-
dihydro-3H-1,2,4-triazole-3-thione) showed excellent antioxidant activity, three times
higher than that of the antibiotic control (cefazolin). 2,20-((((Phenylazanediyl)bis(ethane-
2,1-diyl))bis(4-phenyl-4H-1,2,4-triazole-5,3-diyl))bis(sulfanediyl))diacetate exhibited
notable bactericidal activity against Pseudomonas aeruginosa with MBC and MIC
values 7.3 lg/ml, whereas 12 compounds displayed significant fungicidal activity
against Candida albicans with MIC value 3.9 lg/ml.
Graphical Abstract Synthesis of compounds 2–27. R: 2, 5, 7, 10, 13, 15-27 C6H5–; 3,
8, 11 4-Cl-C6H4–; 4, 9, 12 CH3–; 6, 14 2.6-(CH3)2-C6H3–; R0: 15 CH3CH2–; 16
(CH3)2CH–; 17 (CH3)2CHCH2–; 18 CH3CH2CH2CH2–; 19 (CH3)2CHCH2CH2–; 20
CH3CH2CH2CH2CH2–; 21 NH2COCH2–; 22 C6H5CH2–; 23 C6H5COCH2–; 24
4-ClC6H4COCH2–; 25 4-NO2C6H4COCH2–; 26 C2H5OCOCH2–.
ˇ
Z. J. Beresnevicius
zigmuntas.beresnevicius@ktu.lt
&
1
2
Department of Organic Chemistry, Kaunas University of Technology, 50254 Kaunas, Lithuania
Department of Physical and Inorganic Chemistry, Kaunas University of Technology,
50254 Kaunas, Lithuania
3
Department of Infectious Diseases, Veterinary Academy, Lithuanian University of Health
Sciences, 47181 Kaunas, Lithuania
123

_
I. Tumosiene et al.
H
N
N
S
S
O
O
N
N
N
N
R'
R'
H
N
N
N
R
R
R
NH2
N
N
N
NH2
R
N
N
N
H
S
N
S
N
H
12 -14
15 - 27
1
Keywords Propanehydrazide Á 1,3,4-Oxadiazole Á 1,3,4-Thiadiazole Á
1,2,4-Triazole Á Antioxidative activity Á Antibacterial activity
Introduction
The worsening ecological situation in the world, changing habits of people, and
increasing number of population in large cities influence the weakening human
immunity, emergence of new diseases, spread of new bacteria strains, and mutation
of viruses, etc. Activity of currently used drug preparations decreases or they
become completely inactive. Therefore, search for new biologically active
substances and development of novel pharmaceuticals is an important ongoing
process. Among different five-membered heterocyclic systems, sulfur and nitrogen-
containing ones have gained importance as they constitute the structural features of
many bioactive naturally occurring or synthetic compounds.
Oxadiazoles and thiadiazoles are widely used in medicinal and agricultural
chemistry [1–3]. Substituted 1,3,4-oxadiazole derivatives possess a wide array of
biological activities such as antitubercular [4], antitumor [5], antibacterial and
antifungal [4, 6, 7], anti-convulsant [8], anti-inflammatory [9], and antioxidative [10].
1,3,4-Thiadiazoles are another important class of heterocyclic compounds due to
their numerous biological activities [3] including anticancer [11–13], antimicrobial
and antifungal [14, 15], and antioxidative [16] ones.
1,2,4-Triazole scaffold is yet another important pharmacophore since compounds
bearing this moiety have been shown to display anti-inflammatory and analgesic
[17], antitumor [18], antioxidative [19], antibacterial [13, 14, 20], and antifungal
[21] properties. 1,2,4-Triazole derivatives, e.g., flucarbazone and amicarbazone, are
used as herbicides [22].
Cyclization of hydrazine derivatives is a well-known method for the synthesis of
polyfunctionalized derivatives of 1,3,4-thiadiazole, 1,3,4-oxadiazole, and 1,2,4-
triazole [3, 23, 24].
As a continuation of ongoing efforts on the synthesis of azole derivatives with
potential biological activities and encouraged by the already published data on the
biological activity of 3-arylaminopropanehydrazides and their derivatives [6, 19,
25–27], we report the synthesis of novel derivatives of 3,30-(phenylimino)bis-
propanehydrazide containing two identical heterocyclic moieties and evaluation of
their antioxidative, growth regulating and antimicrobial properties.
123

Synthesis and biological activity of 1,3,4-oxa(thia)diazole…
Experimental
Materials and methods
Chemistry
All the reagents and chemicals were obtained from commercial sources and used
without further purification. Melting points were measured on a B-540 Melting
1
Point Analyzer and are uncorrected. H and 13C NMR spectra were recorded at
ambient temperature on a Varian Unity Inova (300 MHz) and Bruker Avance III/
400 (400 MHz) spectrometers. Chemical shifts (d) are reported in parts per million
1
(ppm) calibrated from TMS (0 ppm) as an internal standard for H NMR, and
DMSO-d6 (39.5 ppm) for 13C NMR. The J constants are given in Hz. IR spectra (m,
cm-1) were recorded on a Perkin–Elmer BX FT–IR spectrometer using KBr pellets.
Elemental analyses were performed on a CE-440 elemental analyzer. TLC was
performed using Silica gel 60 F254 (Kieselgel 60 F254) (Merck, Darmstadt,
Germany) plates.
2,20-(3,30-(Phenylazanediyl)bis(propanoyl))bis(N-phenylhydrazine-1-carboxamide)
(2) Synthesis and characteristics have been described earlier [27].
2,20-(3,30-(Phenylazanediyl)bis(propanoyl))bis(N-(4-chlorophenyl)hydrazine-1-car-
boxamide) (3) To a solution of bispropanehydrazide 1 (1.33 g, 5 mmol) in
methanol (30 ml), 4-chlorophenyl isocyanate (1.54 ml, 1.84 g, 12 mmol) was added
drop-wise and the mixture was heated under reflux for 10 min. The crystalline
precipitate was isolated by filtration and recrystallized from DMF–H2O mixture to
afford 3 as white solid (2.73 g, 95 %), m.p. 218–219 °C; IR (KBr) m/cm-1: 1660
1
(C=O), 3062, 3287, 3308 (NH); H NMR (300 MHz, DMSO-d6) d (ppm): 2.44 (t,
4H, J = 7.2 Hz, CH2CO), 3.58 (t, 4H, J = 7.2 Hz, CH2N), 6.63 (t, 1H, J = 7.2 Hz,
H
(4)Ar), 6.73 (d, 2H, J = 7.2 Hz, H(2,6)Ar), 7.19 (t, 2H, J = 7.2 Hz, H(3,5)Ar), 7.29
(d, 4H, J = 8.85 Hz, H(2,6)Ar0), 7.48 (d, 4H, J = 8.85 Hz, H(3,5)Ar0), 8.14 (s, 2H,
NH), 8.89 (s, 2H, NH), 9.77 (s, 2H, NHAr0). MS (ESI, 20 V) m/z (%): 572
([M?H]?, 30). Anal. Calcd. For C26H27Cl2N7O4 (572.443): C, 54.55; H, 4.75; N,
17.13 %. Found: C, 54.51; H, 4.68; N, 17.09 %.
2,20-(3,30-(Phenylazanediyl)bis(propanoyl))bis(N-methylhydrazine-1-carbothioamide)
(4) To a solution of bispropanehydrazide 1 (1.33 g, 5 mmol) in methanol (30 ml),
methyl isothiocyanate (0.82 ml, 0.88 g, 12 mmol) was added drop-wise and the
reaction mixture was heated under reflux for 20 min. The crystalline precipitate was
isolated by filtration, washed with methanol, and recrystallized from methanol to
afford 4 as white solid (1.60 g, 78 %), m.p. 163–164 °C; IR (KBr) m/cm-1: 1558
1
(C=O), 1682 (C=S), 2986, 3167, 3283 (NH); H NMR (400 MHz, DMSO-d6) d
(ppm): 2.40 (t, 4H, J = 7.2 Hz, COCH2), 2.86 (d, 6H, J = 4.4 Hz, NHCH3), 3.54
(t, 4H, J = 7.2 Hz, CH2N), 6.63 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.73 (d, 2H,
J = 8.4 Hz, H(2,6)Ar), 7.19 (t, 2H, J = 8.4 Hz, H(3,5)Ar), 7.85 (s, 2H, NH), 9.19 (s,
123

_
I. Tumosiene et al.
2H, NH), 9.75 (s, 2H, NHCH3). 13C NMR (100 MHz, DMSO-d6) d (ppm): 30.8
(CH3), 31.1 (CH2CO), 46.1 (CH2N), 111.9, 115.8, 129.2, 146.9 (C-Ar), 170.5
(C=O), 182.1 (C=S). Anal. Calcd. For C16H25N7O2S2 (411.545): C, 46.7; H, 6.12;
N, 23.82 %, Found: C, 46.66; H, 6.09; N, 23.80 %.
2,20-(3,30-(Phenylazanediyl)bis(propanoyl))bis(N-phenylhydrazine-1-carboth-
ioamide) (5) Synthesis and characteristics have been described earlier [27].
2,20-(3,30-(Phenylazanediyl)bis(propanoyl))bis(N-(2,6-dimethylphenyl)hydrazine-1-
carbothioamide) (6) To a solution of bispropanehydrazide 1 (1.33 g, 5 mmol)
dissolved in methanol (30 ml), 2,6-dimethylphenyl isothiocyanate (1.81 ml, 1.96 g,
12 mmol) was added drop-wise and the reaction mixture was heated under reflux for
8 h. Then it was cooled down to room temperature and water was added. The
precipitate was isolated by filtration and recrystallized from DMF–H2O mixture to
afford 6 as white solid (1.63 g, 55 %), m.p. 122–123 °C; IR (KBr) m/cm-1: 1599
1
(C=O), 1679 (C=S), 2972, 3211, 3293 (NH); H NMR (300 MHz, DMSO-d6) d
(ppm): 2.14 (s, 12H, CH3), 2.47 (t, 4H, J = 7.2 Hz, COCH2), 3.59 (t, 4H,
J = 7.2 Hz, CH2N), 6.61 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.73 (t, 2H, J = 8.2 Hz,
H
(2,6)Ar), 6.99–7.12 (m, 6H, HAr0), 7.17 (t, 2H, J = 8.2 Hz, H(3,5)Ar), 9.17 (s, 2H,
NH), 9.46 (s, 2H, NH), 10.02 (s, 2H, NHAr0). 13C NMR (75.4 MHz, DMSO-d6) d
(ppm): 17.9 (4 CH3), 31.5 (CH2CO), 46.1 (CH2N), 111.7, 115.6, 126.8, 127.5,
129.2, 136.6, 136.9, 146.9 (C-Ar), 169.9 (C=O), 181.1 (C=S). Anal. Calcd. For
C30H37N7O2S2 (591.790): C, 60.89; H, 6.30; N, 16.57 %. Found: C, 60.83; H, 6.29;
N, 16.54 %.
N-Phenyl-5-(2-(phenyl(2-(5-(phenylamino)-1,3,4-oxadiazol-2-yl)ethyl)amino)ethyl)-
1,3,4-oxadiazol-2-amine (7) A reaction mixture of bisphenylsemicarbazide 2
(1.51 g, 3 mmol) and POCl3 (10 ml) was heated under reflux for 24 h. The solid
product formed upon pouring the reaction mixture drop-wise into ice–water mixture
(20 ml) was collected by filtration. Filtrate was neutralized with aqueous ammonia;
the crystals were filtered off and washed with water. Combined precipitate was
recrystallized from water to afford 7 as brown solid (0.75 g, 54 %)., m.p.
128–129 °C; IR (KBr) m/cm-1: 1585 (C=O), 1680 (C=S), 2973, 3211, 3287 (NH);
1H NMR (700 MHz, DMSO-d6) d (ppm): 3.02 (t, 4H, J = 6.6 Hz, CH2CN), 3.46 (t,
4H, J = 6.6 Hz, CH2N), 6.58 (t, 1H, J = 7.5 Hz, H(4)Ar), 6.99 (d, 2H, J = 7.5 Hz,
H(2,6) Ar), 7.11 (t, 2H, J = 7.5 Hz, H(3,5) Ar), 7.26–7.64 (m, 10H, HAr0), 10.39 (s,
2H, 2NH). 13C NMR (175 MHz, DMSO-d6) d (ppm): 33.2 (CH2C), 46.4 (CH2N),
112.1, 115.8, 118.3, 121.4, 128.6, 129.9, 139.5, 147.9 (C-Ar), 155.2 (CO), 170.5
(CNH). Anal. Calcd. For C26H25N7O2 (467.5224): C, 66.79; H, 5.39; N, 20.97 %.
Found: C, 66.76; H, 5.35; N, 20.94 %.
N-(4-Chlorophenyl)-5-(2-((2-(5-((4-chlorophenyl)amino)-1,3,4-oxadiazol-2-yl)ethyl)
(phenyl)amino)ethyl)-1,3,4-oxadiazol-2-amine (8) Was prepared from bis-
chlorophenylsemicarbazide 3 (1.72 g, 3 mmol) by following the same synthesis
procedure as for 7 to give 8 as brown solid (0.78 g, 73 %), m.p. 271–272 °C; IR
123

Synthesis and biological activity of 1,3,4-oxa(thia)diazole…
1
(KBr) m/cm-1: 3184 (NH), 1695 (C=N); H NMR (400 MHz, DMSO-d6) d (ppm):
3.09 (t, 4H, J = 6.9 Hz, COCH2), 3.51 (t, 4H, J = 6.9 Hz, CH2N), 6.75–6.89 (m,
5H, HAr), 7.19–7.42 (m, 8H, HAr0), 9.84 (s, 2H, 2NH). 13C NMR (100 MHz,
DMSO-d6), d: 32.7 (CH2C), 47.8 (CH2N), 112.3, 116.2, 119.8, 128.7, 129.5, 138.22,
138.7, 146.8 (C-Ar), 154.90 (CO), 168.80 (CNH). Anal. Calcd. For C26H23Cl2N7O2
(536.413): C, 58.22; H, 4.32; N, 18.28 %. Found: C, 58.18; H, 4.30; N, 18.29 %.
N-Methyl-5-(2-((2-(5-(methylamino)-1,3,4-thiadiazol-2-yl)ethyl)(phenyl)amino)ethyl)-
1,3,4-thiadiazol-2-amine (9) To conc. H2SO4 (5 ml) bismethylthiosemicarbazide 4
(0.82 g, 2 mmol) was added in portions and the reaction mixture was stirred at room
temperature for 15 min. Afterwards, it was poured drop-wise into ice–water mixture
and neutralized with Na2CO3. The crystalline precipitate was isolated by filtration,
washed with water and recrystallized from methanol–water mixture to afford 9 as
light brown solid (0.49 g, 65 %), m.p. 167–168 °C; IR (KBr) m/cm-1: 1598 (C=N),
1
3190 (NH); H NMR (400 MHz, DMSO-d6) d (ppm): 3.04 (t, 4H, J = 7.0 Hz,
COCH2), 2.86 (d, 6H, J = 4.9 Hz, NHCH3), 3.63 (t, 4H, J = 7.0 Hz, CH2N), 6.68
(t, 1H, J = 7.0 Hz, H(4)Ar), 6.76 (d, 2H, J = 8.4 Hz, H(2,6)Ar), 7.22 (t, 2H,
J = 8.4 Hz, H(3,5)Ar), 7.51 (q, 2H, NHCH3). 13C NMR (100 MHz, DMSO-d6) d
(ppm): 27.3 (CH3), 31.1 (CH2C), 49.8 (CH2N), 112.3, 116.3, 129.3, 146.6, 154.8
(CS), 169.4 (CNH). Anal. Calcd. For C16H21N7S2 (375.515): C, 51.18; H, 5.64; N,
26.11 %, Found: C, 51.16; H, 5.65; N, 26.09 %.
N-Phenyl-5-(2-(phenyl(2-(5-(phenylamino)-1,3,4-thiadiazol-2-yl)ethyl)amino)ethyl)-
1,3,4-thiadiazol-2-amine (10) Was prepared from bisphenylthiosemicarbazide 5
(1.61 g, 3 mmol) by following the same synthesis procedure as for 9 to give 10 as
white solid (1.18 g, 47 %), m.p. 154–155 °C; IR (KBr) m/cm-1: 1600 (C=N), 3196
1
(NH); H NMR (300 MHz, DMSO-d6) d (ppm): 3.15 (t, 4H, J = 6.6 Hz, CH2C),
3.72 (t, 4H, J = 6.6 Hz, CH2N), 6.71 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.79–6.89 (m,
2H, H(2,6)Ar), 6.98 (t, 2H, J = 7.2 Hz, H(3,5)Ar), 7.24 (t, 2H, J = 8.0 Hz, H(4)Ar0),
7.32 (t, 4H, J = 8.0 Hz, H(3,5)Ar0), 7.59 (d, 4H, J = 8.0 Hz, H(2,6)Ar0), 10.32 (s, 2H,
2NH). 13C NMR (75.4 MHz, DMSO-d6) d (ppm): 27.1 (CH2C), 49.8 (CH2N),
112.9, 115.4, 117.2, 121.7, 129.0, 129.4, 140.6, 146.3 (C-Ar), 156.8 (CS), 164.4
(CNH). Anal. Calcd. For C26H25N7S2 (499.654): C, 62.50, H, 5.04; N, 19.62 %.
Found: C, 62.45; H, 5.01; N, 19.60 %.
5,50-((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-(4-chlorophenyl)-2,4-dihydro-3H-
1,2,4-triazol-3-one) (11) A mixture of bischlorophenylsemicarbazide 3 (1.72 g,
3 mmol) and 20 % aqueous KOH solution (25 ml) was heated under reflux for 4 h
and cooled to room temperature. Afterwards conc. HCl was added to pH 4. The
crystalline precipitate was isolated by filtration, washed with water, and recrystal-
lized from DMF–H2O mixture to afford 11 as white solid (0.98 g, 61 %), m.p.
1
314–315 °C; IR (KBr) m/cm-1: 1588 (C=N), 1707 (C=O), 3172 (NH); H NMR
(300 MHz, DMSO-d6) d (ppm): 2.56 (t, 4H, J = 6.9 Hz, CH2CN), 3.30 (t, 4H,
J = 6.9 Hz; CH2N), 6.58 (t, 1H, J = 8.7 Hz, H(4)Ar), 7.39 (d, 2H, J = 8.7 Hz,
H
(3,5)Ar), 7.42 (d, 2H, J = 8.7 Hz, H(2,6)Ar), 7.55 (d, 4H, J = 8.7 Hz, H(3,5)Ar0),
123

_
I. Tumosiene et al.
7.58 (d, 4H, J = 8.7 Hz, H(2,6)Ar0), 11.82 (s, 2H, NNH). MS (ESI, 20 V) m/z (%):
537 ([M?H]?, 40). Anal. Calcd. For C26H23Cl2N7O2 (536.413): C, 58.22; H, 4.32;
N, 18.28 %. Found: C, 58.19; H, 4.30; N, 18.17 %.
5,50-((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-methyl-2,4-dihydro-3H-1,2,4-tria-
zole-3-thione) (12) Was prepared from bismethylthiosemicarbazide 4 (1.24 g,
3 mmol) by following the same synthesis procedure as for 11 to give 12 as white
solid (1.02 g, 90 %), m.p. 275–276 °C; IR (KBr) m/cm-1: 1339 (C=S), 1599 (C=N),
1
3093 (NH); H NMR (400 MHz, DMSO-d6) d (ppm): 2.89 (t, 4H, J = 7.2 Hz,
COCH2), 3.38 (s, 6H, CH3), 3.68 (t, 4H, J = 7.2 Hz, CH2N), 6.64 (t, 1H,
J = 7.2 Hz, H(4)Ar), 6.72 (d, 2H, J = 8.0 Hz, H(2,6)Ar), 7.18 (t, 2H, J = 8.4 Hz,
H
(3,5)Ar), 13.57 (s, 2H, NH). 13C NMR (100 MHz, DMSO-d6) d (ppm): 22.9 (CH3-
N); 29.7 (CH2C); 46.7 (CH2N); 111.7, 116.2, 129.4, 146.5 (C-Ar), 150.9 (CN);
166.5 (C=S). Anal. Calcd. For C16H21N7S2 (375.515): C, 51.18; H, 5.64; N,
26.11 %. Found: C, 51.10; H, 5.60; N, 26.09 %.
5,50-((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-2,4-dihydro-3H-1,2,4-tria-
zole-3-thione) (13) Synthesis and characteristics have been described earlier [27].
5,50-((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-(2,6-dimethylphenyl)-2,4-dihydro-
3H-1,2,4-triazole-3-thione) (14) Was prepared from bisdimethylthiosemicar-
bazide 6 (1.77 g, 3 mmol) by following the same synthesis procedure as for 11,
except that the reaction duration was 1 h, to afford 14 as white solid (1.48 g, 89 %),
m.p. 160–161 °C; IR (KBr) m/cm-1: 1291 (C=S), 1600 (C=N), 3085 (NH); 1H NMR
(400 MHz, DMSO-d6) d (ppm): 1.92 (s, 12H, CH3), 2.35–2.56 (m, 4H, CH2CN),
3.40–3.61 (m, 4H, CH2N), 6.59–7.44 (m, 11H, HAr?Ar0),14.00 (s, 2H, NNH). 13C
NMR (100 MHz, DMSO-d6) d (ppm): 17.3 (4 CH3), 31.1 (CH2C), 40.4 (CH2N),
111.8, 117.1, 128.8, 129.4, 129.7, 135.7, 136.0, 146.5 (C-Ar), 155.2 (CN), 166.6
(C=S). Anal. Calcd. For C30H33N7S2 (555.760): C, 64.83; H, 5.98; N, 17.64 %.
Found: C, 64.79; H, 5.95; N, 17.61 %.
General procedure for synthesis of N,N-bis({2-[5-substituted thio)-4-phenyl-4H-
1,2,4-triazol-3-yl]ethyl})anilines 15–26 Method A. To a solution of bistriazo-
lethione 13 (0.40 g, 0.8 mmol) in DMF (5 ml), NaH (0.048 g, 2 mmol) was added,
the reaction mixture was stirred at room temperature until evolution of hydrogen
stopped (approx. 10 min), and haloalkane (2 mmol) was added drop-wise. The
reaction mixture was stirred at 50 °C for 20 h. Afterwards, cold water (20 ml) was
added, the precipitate formed was isolated by filtration, and recrystallized from
2-propanol.
Method B. To a solution of bistriazolethione 13 (0.40 g, 0.8 mmol) in DMF
(5 ml), KOH powder (0.11 g, 2 mmol), K2CO3 (0.3 g, 2,2 mmol), and haloalkane
(2 mmol) were added. The reaction mixture was stirred at 35–40 °C for 24 h.
Afterwards cold water (30 ml) was added, the precipitate formed was isolated by
filtration, and recrystallized from 2-propanol (unless indicated otherwise).
123

Synthesis and biological activity of 1,3,4-oxa(thia)diazole…
Method C. To a solution of 13 (0.40 g, 0.8 mmol) in DMF (5 ml), triethylamine
(0.20 g, 0.28 ml, 2 mmol) and haloalkane (2 mmol) were added. The reaction
mixture was stirred at room temperature for 4 h. Afterwards cold water (30 ml) was
added, the precipitate formed was isolated by filtration, and recrystallized from
2-propanol.
N,N-Bis(2-(5-(ethylthio)-4-phenyl-4H-1,2,4-triazol-3-yl)ethyl)aniline
(15) Pre-
pared according to method A from bistriazolethione 13 and iodoethane (0.31 g,
0.16 ml). White solid (0.24 g, 53 %), m.p. 195–196 °C; IR (KBr) m/cm-1: 1596
1
(C=N); H NMR (400 MHz, DMSO-d6) d (ppm): 1.25 (t, 6H, J = 7.2 Hz, CH3),
2.11 (s, 3H, CH3), 2.68 (t, 4H, J = 7.2 Hz, CH2C), 3.04 (q, 4H, J = 7.2 Hz,
J = 14.8 Hz, SCH2CH3), 3.35 (t, 4H, J = 7.2 Hz, NCH2), 5.94 (d, 2H, J = 8.4 Hz,
H
(2,6)Ar), 6.48 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.87 (t, 2H, J = 7.2 Hz, H(3,5)Ar),
7.36–7.41 (m, 4H, H(3,5)Ar0), 7.53–7.60 (m, 6H, H(2,4,6)Ar0). 13C NMR (100 MHz,
DMSO-d6) d (ppm): 14.8 (CH3), 22.5 (CH2C), 26.5 (CH2), 47.6 (CH2N), 110.8,
115.7, 127.4, 129.0, 129.9, 129.9, 132.9, 145.9 (C-Ar), 149.8 (CN), 153.5 (CS).
Anal. Calcd. For C30H33N7S2 (555.760): C, 64.83; H, 5.98; N, 17.64 %. Found: C,
64.53; H, 6.03; N, 17.60 %.
N,N-Bis(2-(5-(isopropylthio)-4-phenyl-4H-1,2,4-triazol-3-yl)ethyl)aniline (16) Pre-
pared according to method A from bistriazolethione 13 and 2-iodopropane
(0.34 g, 0.20 ml). White solid (0.26 g, 56 %), m.p. 185–186 °C; IR (KBr) m/
cm-1: 1597 (C=N); 1H NMR (400 MHz, DMSO-d6) d (ppm): 1.25 (d, 12H, 2 CH3,
J = 6.8 Hz), 2.11 (s, 3H, CH3), 2.69 (t, 4H, J = 7.6 Hz, CH2C), 3.34 (t, 4H,
J = 7.2 Hz, NCH2), 3.57 (sep, 2H, J = 6.8 Hz, CH), 5.94 (d, 2H, J = 7.8 Hz,
H
(2,6)Ar), 6.48 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.87 (t, 2H, J = 7.8 Hz, H(3,5)Ar),
7.33–7.40 (m, 4H, H(3,5)Ar0), 7.53–7.61 (m, 6H, H(2,4,6)Ar0). 13C NMR (100 MHz,
DMSO-d6) d (ppm): 22.6 (CH2C), 23.0 (CH3), 38.4 (CH), 47.6 (CH2N), 110.8,
115.7, 127.5, 129.0, 129.8, 129.9, 133.0, 145.9 (C-Ar), 149.2 (CN), 153.5 (CS).
Anal. Calcd. For C32H37N7S2 (583.813): C, 65.83; H, 6.39; N, 16.79 %, Found: C,
65.74; H, 6.32; N, 16.75 %.
N,N-Bis(2-(5-(isobutylthio)-4-phenyl-4H-1,2,4-triazol-3-yl)ethyl)aniline (17) Pre-
pared according to method A from bistriazolethione 13 and 1-bromo-2-methyl-
propane (0.34 g, 0.20 ml). White solid (0.24 g, 50 %), m.p. 154–155 °C; IR (KBr)
1
m/cm-1: 1597 (C=N); H NMR (400 MHz, DMSO-d6) d (ppm): 0.91 (d, 12H,
J = 6.4 Hz, CH3), 1.86 (sep, 2H, J = 6.8 Hz, CH), 2.69 (t, 4H, J = 7.4 Hz, CH2-
C), 2.96 (d, 4H, J = 6.8 Hz, SCH2), 3.36 (t, 4H, J = 7.4 Hz, NCH2), 5.95 (d, 2H,
J = 8.0 Hz, H(2,6)Ar), 6.50 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.89 (t, 2H, J = 7.8 Hz,
H
(3,5)Ar), 7.35–7.41 (m,4H, H(3,5)Ar0), 7.56–7.63 (m, 6H, H(2,4,6)Ar0). 13C NMR
(100 MHz, DMSO-d6) d (ppm): 21.3 (CH3), 22.5 (CH2C), 27.8 (CH), 40.5 (CH2),
47.6 (CH2N), 110.7, 115.6, 127.4, 129.0, 129.9, 129.9, 132.9, 145.9 (C-Ar), 150.0
(CN), 153.5 (CS). Anal. Calcd. For C34H41N7S2 (611.866): C, 66.74; H, 6.75; N,
16.02 %. Found: C, 66.71; H, 6.70; N, 15.97 %.
123

_
I. Tumosiene et al.
N,N-Bis(2-(5-(butylthio)-4-phenyl-4H-1,2,4-triazol-3-yl)ethyl)aniline
(18) Pre-
pared according to Method A from bistriazolethione 13 and 1-iodobutane (0.34 g,
0.23 ml). White solid (0.25 g, 57 %), m.p. 145–146 °C; IR (KBr) m/cm-1: 1597
1
(C=N), 2929 (NC); H NMR (400 MHz, DMSO-d6) d (ppm): 0.84 (t, 6H, CH3,
J = 7.2 Hz), 1.31 (sxt, 4H, CH2, J = 7.6 Hz), 1.57 (qui, 4H, SCH2CH2,
J = 7.6 Hz), 2.67 (t, 4H, J = 7.6 Hz, CH2C), 3.03 (t, 4H, J = 7.2 Hz, SCH2),
3.34 (t, 4H, J = 7.2 Hz, NCH2), 5.94 (d, 2H, J = 8.4 Hz, H(2,6)Ar), 6.48 (t, 1H,
J = 7.2 Hz, H(4)Ar), 6.87 (t, 2H, J = 7.2 Hz, H(3,5)Ar), 7.32–7.42 (m, 4H,
H
(3,5)Ar0), 7.51–7.63 (m, 6H, H(2,4,6)Ar0). 13C NMR (100 MHz, DMSO-d6) d (ppm):
13.3 (CH3), 20.9 (CH2), 22.5 (CH2C), 30.9 (CH2), 31.8 (CH2), 47.6 (CH2N), 110.8,
115.7, 127.4, 129.0, 129.9, 129.9, 132.9, 145.9 (C-Ar), 149.9 (CN), 153.5 (CS).
Anal. Calcd. For C34H41N7S2 (611.866): C, 66.74; H, 6.75; N, 16.02 %. Found: C,
66.68; H, 6.69; N, 15.97 %.
N,N-Bis(2-(5-(isopentylthio)-4-phenyl-4H-1,2,4-triazol-3-yl)ethyl)aniline (19) Pre-
pared according to method A from bistriazolethione 13 and 1-iodo-3-methylbutane
(0.40 g, 0.26 ml). White solid (0.27 g, 53 %), m.p. 153–154 °C; IR (KBr) m/cm-1
:
1597 (C=N); 1H NMR (400 MHz, DMSO-d6) d (ppm): 0.84 (d, 12H, J = 6.8 Hz, 2
CH3), 1.48 (q, 4H, J = 7.2 Hz, J = 14.4 Hz, CH2),1.58 (sep, 2H, J = 6.6 Hz, CH),
2.67 (t, 4H, J = 7.2 Hz, CH2C), 3.03 (t, 4H, J = 7.2 Hz, SCH2), 3.33 (t, 4H,
J = 7.2 Hz, NCH2), 5.94 (d, 2H, J = 8.0 Hz, H(2,6)Ar), 6.48 (t, 1H, J = 7.2 Hz,
H
(4)Ar), 6.87 (t, 2H, J = 7.2 Hz, H(3,5)Ar), 7.32–7.43 (m, 4H, H(3,5)Ar0), 7.51–7.63
(m, 6H, H(2,4,6)Ar0). 13C NMR (100 MHz, DMSO-d6) d (ppm): 21.9 (CH3), 22.5
(CH2N), 26.6 (CH), 30.2 (CH2), 37.8 (CH2), 47.6 (CH2N), 110.8, 115.7, 127.4,
129.0, 129.9, 129.9, 132.9, 145.9 (C-Ar), 149.9 (CN), 153.6 (CS). Anal. Calcd. For
C36H45N7S2 (639.919): C, 67.57; H, 7.09; N, 15.32 %. Found: C, 67.49; H, 7.11; N,
15.29 %.
N,N-Bis(2-(5-(pentylthio)-4-phenyl-4H-1,2,4-triazol-3-yl)ethyl)aniline (20) Pre-
pared according to method A from bistriazolethione 13 and 1-iodopentane
(0.40 g, 0.26 ml). White solid (0.18 g, 70 %), m.p. 128–129 °C; IR (KBr) m/
cm-1: 1597 (C=N); 1H NMR (400 MHz, DMSO-d6) d (ppm): 0.85 (t, 6H,
J = 7.2 Hz, CH3), 1.22–1.32 (m, 8H, CH2CH2), 1.61 (qui, 4H, J = 7.2 Hz,
SCH2CH2), 2.69 (t, 4H, J = 7.2 Hz, CH2-C), 3.04 (t, 4H, J = 7.2 Hz, SCH2), 3.37
(t, 4H, J = 7.2 Hz, NCH2), 5.98 (d, 2H, J = 7.2 Hz, H(2,6)Ar), 6.51 (t, 1H,
J = 7.2 Hz, H(4)Ar), 6.90 (d, 2H, J = 8.1 Hz, H(3,5)Ar), 7.37–7.41 (m, 4H,
H
(3,5)Ar0), 7.54–7.63 (m, 6H, H(2,4,6)Ar0). 13C NMR (175 MHz, DMSO-d6) d (ppm):
14.3 (CH3), 22.1 (CH2C), 23.1 (CH2), 29.1 (CH2), 30.5 (CH2), 32.6 (CH2), 48.1
(CH2N), 111.3, 116.2, 127.9, 129.6, 130.4, 130.4, 133.5, 146.5 (C-Ar), 150.4 (CN),
154.06 (CS). Anal. Calcd. For C36H45N7S2 (639.919): C, 67.57; H, 7.09; N,
15.32 %. Found: C, 67.54; H, 7.05; N, 15.29 %.
2,20-((((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-4H-1,2,4-triazole-5,3-diyl))
bis(sulfanediyl))diacetamide (21) Prepared according to method B from bistriazo-
lethione 13 and 2-chloroacetamide (0.19 g). White solid (0.41 g, 84 %), m.p.
123

Synthesis and biological activity of 1,3,4-oxa(thia)diazole…
1
68–69 °C; IR (KBr) m/cm-1: 1598 (C=N), 1679 (C=O), 3322 (NH2); H NMR
(400 MHz, DMSO-d6) d (ppm): 2.70 (t, 4H, J = 7.2 Hz, CH2C), 3.39 (t, 4H,
J = 7.2 Hz, NCH2), 3.89 (s, 4H, SCH2), 6.01 (d, 2H, J = 8.0 Hz, H(2,6)Ar), 6.52 (t,
1H, J = 7.2 Hz, H(4)Ar), 6.91 (t, 2H, J = 7.2 Hz, H(3,5)Ar), 7.42–7.45 (m, 4H,
H
(3,5)Ar0), 7.59–7.63 (m, 6H, H(2,4,6)Ar0), 7.66 (s, 4H. NH2). 13C NMR (100 MHz,
DMSO-d6) d (ppm): 22.4 (CH2C), 35.9 (CH2), 47.7 (CH2N), 110.9, 115.7, 127.2,
129.0, 129.9, 129.9, 132.7, 145.9 (C-Ar), 149.8 (CN), 168.5 (CS), 194.2 (C=O).
Anal. Calcd. For C30H31N9O2S2 (613.756): C, 58.71; H, 5.09; N, 20.54 %. Found:
C, 58.67; H, 5.03; N, 20.51 %.
N,N-Bis(2-(5-(benzylthio)-4-phenyl-4H-1,2,4-triazol-3-yl)ethyl)aniline (22) Pre-
pared according to method B from bistriazolethione 13 and chlorophenylmethane
(0.25 g). White solid (0.38 g, 69 %), m.p. 172–173 °C; IR (KBr) m/cm-1: 1597
1
(C=N); H NMR (400 MHz, DMSO-d6) d (ppm): 2.68 (t, 4H, J = 7.2 Hz, CH2C),
3.35 (t, 4H, J = 7.2 Hz, CH2N), 4.33 (s, 4H, SCH2), 5.94 (d, 2H, J = 8.4 Hz,
H
(2,6)Ar), 6.88 (d, 2H, J = 8.4 Hz, H(3,5)Ar), 7.23–7.33 (m, 12H, HAr0 ? HAr00),
7.51–7.60 (m, 8H, HAr0). 13C NMR (100 MHz, DMSO-d6) d (ppm): 22.5 (CH2C),
36.3 (CH2), 47.7 (CH2N), 110.8, 115.7, 128.4, 128.9, 127.4, 129.7, 129.8, 129.9,
132.8, 134.4, 137.1, 145.8 (C-Ar), 149.5 (CN), 153.7 (CS). Anal. Calcd. For
C40H37N7S2 (679.899): C, 70.66; H, 5.49; N, 14.42 %. Found: C, 70.61; H, 5.45; N,
14.41 %.
2,20-((((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-4H-1,2,4-triazole-5,3-diyl))
bis(sulfanediyl))bis(1-phenylethan-1-one) (23) Prepared according to method C
from bistriazolethione 13 and 2-bromo-1-phenylethanone (0.40 g). White solid
(0.40 g, 68 %), m.p. 84–85 °C; IR (KBr) m/cm-1: 1597 (C=N), 1679 (C=O), 2915; 1
H NMR (700 MHz, DMSO-d6) d (ppm): 2.70 (t, 4H, J = 7.2 Hz, CH2C), 3.37 (t,
4H, J = 7.2 Hz, CH2N), 4.88 (s, 4H, SCH2), 6.01 (d, 2H, J = 6.4 Hz, H(2,6)Ar),
6.52 (t, 1H, J = 6.4 Hz, H(4)Ar), 6.92 (t, 2H, J = 6.4 Hz, H(3,5)Ar), 7.43–7.70 (m,
16H, HAr0 ? HAr00), 8.02 (d, 4H, J = 6.4 Hz, HAr00). 13C NMR (175 MHz,
DMSO-d6) d (ppm): 22.4 (CH2C), 40.1 (CH2), 47.7 (CH2N), 110.8, 115.7, 127.3,
128.3, 128.7, 129.04, 129.9, 130.0, 132.7, 133.7, 135.2, 145.9 (C-Ar), 149.4 (CN),
153.6 (CS), 193.0 (C=O). Anal. Calcd. For C42H37N7O2S2 (735.919): C, 68.55; H,
5.07; N, 13.32 %. Found: C, 68.49; H, 5.03; N, 13.28 %.
2,20-((((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-4H-1,2,4-triazole-5,3-diyl))
bis(sulfanediyl))bis(1-(4-chlorophenyl)ethan-1-one) (24) Prepared according to
method C from bistriazolethione 13 and 2-bromo-1-(4-chlorophenyl)ethanone
(0.47 g). White solid (0.51 g, 79 %), m.p. 171–172 °C; IR (KBr) m/cm-1: 1589
1
(C=N), 1681 (C=O); H NMR (700 MHz, DMSO-d6) d (ppm): 2.69 (t, 4H,
J = 7.2 Hz, CH2C), 3.38 (t, 4H, J = 7.2 Hz, CH2N), 4.85 (s, 4H, SCH2), 6.00 (d,
2H, J = 7.2 Hz, H(2,6)Ar), 6.51 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.91 (t, 2H, J = 7.2 Hz,
H
(3,5)Ar), 7.42 (dd, 4H, J = 1.6 Hz, J = 6.0 Hz, H(3,5)Ar0), 7.56–7.67 (m, 10H,
HAr0 ? HAr00), 8.03 (d, 4H, J = 6.4 Hz, HAr00). 13C NMR (175 MHz, DMSO-d6) d
(ppm): 22.4 (CH2C), 40.0 (CH2), 47.66 (CH2N), 110.8, 115.7, 127.2, 128.9, 129.0,
123

_
I. Tumosiene et al.
129.9, 130.0, 130.3, 132.7, 133.9, 138.6, 145.9 (C-Ar), 149.3 (CN), 153.7 (CS),
192.2 (C=O). Anal. Calcd. For C42H35Cll2N7O2S2 (804.809): C, 62.68; H, 4.38; N,
12.18 %. Found: C, 62.63; H, 4.31; N, 12.15 %.
2,20-((((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-4H-1,2,4-triazole-5,3-
diyl))bis(sulfanediyl))bis(1-(4-nitrophenyl)ethan-1-one) (25) Prepared accord-
ing to Method C from bistriazolethione 13 and 2-bromo-1-(4-nitrophenyl)ethanone
(0.49 g). Brown solid (0.60 g, 90 %), m.p. 150–151 °C; IR (KBr) m/cm-1: 1345
(NO2), 1589 (C=N), 1681 (C=O); 1H NMR (400 MHz, DMSO-d6) d (ppm): 2.59 (t,
4H, J = 7.2 Hz, CH2C), 3.38 (t, 4H, J = 7.2 Hz, CH2N), 5.26 (s, 4H, SCH2), 6.02
(d, 2H, J = 7.2 Hz, H(2,6)Ar,), 6.53 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.93 (t, 2H,
J = 7.2 Hz, H(3,5)Ar), 7.38–7.43 (m, 4H, H(3,5)Ar0), 7.55–7.59 (m, 6H, H(2,4,6)Ar0),
8.22–8.46 (m, 8H, HAr00). 13C NMR (175 MHz, DMSO-d6) d (ppm): 23.1 (CH2C),
30.5 (CH2), 46.9 (CH2N), 110.8, 115.9, 123.8, 128.3, 129.0, 129.7, 130.0, 130.3,
132.8, 133.5, 140.1, 145.7 (C-Ar), 150.1 (CN), 153.9 (CS), 193.9 (C=O). Anal.
Calcd. For C42H35N9O6S2 (825.914): C, 61.08; H, 4.27; N, 15.26 %. Found: C,
61.01; H, 4.22; N, 15.27 %.
Diethyl
2,20-((((phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-4H-1,2,4-tria-
zole-5,3-diyl))bis(sulfanediyl))diacetate (26) Prepared according to Method B
from bistriazolethione 13 and 2.45 g (2.14 ml) ethyl chloroacetate (20 mmol).
White solid (0.48 g, 89 %), m.p. 153–154 °C; IR (KBr) m/cm-1: 1597 (C=N), 1730
1
(C=O); H NMR (400 MHz, DMSO-d6) d (ppm): 1.19 (t, 6H, J = 7.2 Hz, CH3),
2.70 (t, 4H, J = 7.2 Hz, CH2C), 3.08 (t, 4H, J = 7.2 Hz, NCH2), 4.03 (s, 4H,
SCH2), 4.11 (q, 4H, J = 7.2 Hz, J = 14.0 Hz, OCH2CH3), 5.98 (d, 2H,
J = 8.0 Hz, H(2,6)Ar), 6.51 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.90 (t, 2H, J = 7.2 Hz
H
(3,5)Ar), 7.41–7.43 (m, 4H, H(3,5)Ar0), 7.59–7.62 (m, 6H, H(2,4,6)Ar0). 13C NMR
(100 MHz, DMSO-d6) d (ppm): 13.9 (OCH2CH3), 22.4 (CH2C), 33.9 (CH2), 47.6
(CH2N), 61.2 (OCH2CH3), 110.9, 115.8, 127.2, 129.1, 130.0, 130.1, 132.6, 145.87
(C-Ar), 149.2 (CN), 153.8 (C=O), 168.0 (CS). Anal. Calcd. For C34H37N7O4S2
(671.832): C, 60.78; H, 5.55; N, 14.59 %, Found: C, 60.72; H, 5.51; N, 14.56 %.
2,20-((((Phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-4H-1,2,4-triazole-5,3-
diyl))bis(sulfanediyl))di(acetohydrazide) (27) To a solution of bisphenyltriazole-
sulfaneylacetate 26 (5.37 g, 8 mmol) in methanol (100 ml), hydrazine hydrate
(0.62 ml, 0.64 g, 20 mmol) was added and the reaction mixture was stirred at room
temperature for 12 h. Afterwards H2O (50 ml) was added and the mixture was left
to stand at 4 °C for 24 h. The precipitate was isolated by filtration and recrystallized
from 2-propanol to afford 27 as white solid (3.59 g, 70 %), m.p. 105–106 °C; IR
(KBr) m/cm-1: 1598 (C=N), 1674 (C=O), 3051, 3252 (NH, NH2); 1H NMR
(400 MHz, DMSO-d6) d (ppm): 2.71 (t, 4H, J = 7.2 Hz, CH2C), 3.39 (t, 4H,
J = 7.2 Hz, NCH2), 3.86 (s, 4H, SCH2), 4.28 (br.s, 4H, NH2), 5.98 (d, 2H,
J = 7.2 Hz, H(2,6)Ar), 6.51 (t, 1H, J = 7.2 Hz, H(4)Ar), 6.90 (t, 2H, J = 7.2 Hz
H
(3,5)Ar), 7.38–7.44 (m, 4H, H(3,5)Ar0), 7.56–7.64 (m, 6H, H(2,4,6)Ar0), 9.37 (s, 2H,
NH). 13C NMR (100 MHz, DMSO-d6) d (ppm): 22.5 (CH2C), 34.2 (CH2), 47.7
123

Synthesis and biological activity of 1,3,4-oxa(thia)diazole…
(CH2N), 110.9, 115.8, 127.3, 129.1, 129.9, 130.1, 132.7, 145.9 (C-Ar), 149.7 (CN),
153.8 (C=O), 168.0 (CS). Anal. Calcd. For C30H33N11O2S2 (643.786): C, 55.97; H,
5.17; N, 23.93 %, Found: C, 55.89; H, 5.15; N, 23.90 %.
Biology
Evaluation of antioxidative activity Free radical scavenging activity of compounds
was measured by DPPH using the widely used method [28]. 1 mM solution of
DPPH in ethanol was prepared, and this solution (1 ml) was added to the solutions
of tested compounds (1 mg ml-1 of DMSO). The mixture was shaken vigorously
and allowed to stand at room temperature for 20 min. The absorbance was measured
at 517 nm with a spectrophotometer (UV-200-RS). Lower absorbance of the
reaction mixture indicated higher free radical scavenging activity. The capability to
scavenge the DPPH radical was calculated according to the following equation:
DPPH scavenging effect ð%Þ ¼ ðA0 À A1=A0Þ Â 100;
where A0 is the absorbance of the control reaction and A1 is the absorbance in the
presence of the samples.
Evaluation of reducing power Compounds 2–27 (1 mg) were dissolved in 1 ml of
DMSO. The obtained solutions were mixed with 2.5 ml of (pH 6.6) 0.2 M
phosphate buffer and 2.5 ml of 1 % potassium ferricyanide. The mixture was
incubated at 50 °C for 20 min. 2.5 ml of 10 % trichloroacetic acid were added to
the mixture, which was then centrifuged for 10 min at 1000 rpm. 2.5 ml of the
upper layer of the solution was mixed with 2.5 ml of distilled water and 0.5 ml of
0.1 % ferric chloride. The absorbance was measured at 700 nm with a spectropho-
tometer (UV-200-RS) [29].
Rapeseed organogenesis in vitro Seeds of rapeseed (Brassica napus L.) ‘Terra’
were used for the experiments. Seeds were initially washed in ethanol for 1 min;
afterwards they were washed in bleach for 20 min and rinsed with distilled water.
After sterilization, seeds were placed in 9-cm Petri dishes (total amount 150) on filter
paper, which was wetted with 3 ml of distilled water (control sample) or solution of
azole derivative. Azoles were examined at 0.5 and 1.0 mg l-1 concentrations. The
results of experiments were compared with the ones of the control sample. Each Petri
dish contained ten seeds and was sealed with a strip of parafilm to prevent
evaporation. Seeds were germinated in the dark for 7 days at room temperature
(22 °C) [30]. After 7 days number of leaves was counted, height of seedlings and
length of roots were measured. Each experiment was repeated three times.
Biometric measurements The length of axial organs of 30 selected rapeseed
seedlings per treatment was measured. In order to assess the tolerance of seedlings
towards azoles we applied
a Wilkinson tolerance index (WTI): It =
(lme/lc) 9 100 %, where lme indicates the increase in axial organ growth in an
azole solution and lc is the increase in axial organ growth in the control sample [31].
123

_
I. Tumosiene et al.
Evaluation of antimicrobial activity The following bacteria strains were used:
Gram-positive cocci Staphylococcus aureus (ATCC 9144) Gram-negative rods
Escherichia coli (ATCC 8739), Pseudomonas aeruginosa (NCTC 6750) and fungi
Candida albicans (ATCC 10,231). Tryptic soy agar (TSA) and tryptic soy broth
(TSB) were used for bacteria cultivation and antibacterial activity tests. Antimi-
crobial activity of the compounds was determined by testing their different
concentrations against B. cereus, S. aureus, E. coli, P. aeruginosa and C. albicans
strains by the broth-dilution and spread plate methods [32, 33]. The test bacteria
S.aureus, E.coli, P. aeruginosa and fungi C. albicans were streaked out on TSA
plates and incubated at 37 °C for 24 h. A representative colony was placed in 5 ml
of TSB and incubated at 37° C for 24 h. S. aureus, E.coli, P. aeruginosa and
C.albicans cultures containing 107 CFU/ml (colony-forming units, corresponding to
MC Farland’s) were prepared by dilution with TSB and used for antimicrobial tests.
Solutions of tested compounds in a range of concentrations, 1000, 500, 250, 125,
62.5, 31.25, 15.6, 7.3, and 3.9 lg/ml, were prepared for each sample. The test
organisms (100 lL) were added to each tube and incubated at 37 °C for 24 h. At the
end of this period, a small amount of the diluted mixture from each tube was pulled
out and spread on TSA. The plates were incubated at 37 °C for 48 h. The growth of
bacterial cells was observed on agar plates. The lowest concentration of the bacterial
material at which no growth was observed was considered as the minimum
bactericidal concentration (MBC) value [34]. Minimum inhibitory concentration
(MIC) is the lowest concentration of an antimicrobial that inhibits the visible growth
of a microorganism after overnight incubation. Oxytetracycline inoculated with test
bacteria S. aureus, E. coli, and P. aeruginosa in the tubes and plates was used as a
control for antibacterial activity screening. Nystatine inoculated with test fungi C.
albicans in the tubes and plates was used as a control for antifungal activity
screening. The growth of the test microorganism cells was observed on agar plates.
Results and discussion
Chemistry
Reaction of bispropanehydrazide 1 with phenyl or 4-chlorophenyl isocyanates, or
methyl, phenyl or 2,6-dimethylphenyl isothiocyanate in methanol gave bishy-
drazinecarboxamides 2 and 3, and bishydrazinecarbothiamide derivatives 4–6
(Scheme 1). In the reaction of bispropanehydrazide 1 with phenyl isocyanates,
crystals of the bisphenylsemicarbazides 2–3 began to appear already after heating
for 10 min, whereas for the reactions with phenyl isothiocyanates more prolonged
heating was required. The 1H NMR spectra of bisphenylsemicarbazides 2 and 3 and
bisthiosemicarbazides 4–6 show a set of resonances for the protons attributable to
the phenyl moiety, whereas the signals of the primary amino group present in the
bispropanehydrazide 1 are absent.
Hydrazinecarboxamides undergo cyclization in phosphoryl chloride or sulfuric
acid, forming 1,3,4-oxadiazole derivatives. N-Phenyl-5-(2-(phenyl(2-(5-(pheny-
lamino)-1,3,4-oxadiazol-2-yl)ethyl)amino)ethyl)-1,3,4-oxadiazol-2-amine (7), and
123

Synthesis and biological activity of 1,3,4-oxa(thia)diazole…
S
O
H
H
N
O
O
O
O
N
H
N
NH2
N
H
N
H
N
H
R
R
N
H
R
R
a
b
N
N
e
N
S
O
H
N
H
N
NH2
N
H
N
H
N
H
N
H
N
H
O
O
2, 3
1
4 - 6
c
d
H
H
N
H
N
N
N
N
N
O
N
N
R
R
R
S
N
O
O
R
R
N
N
N
S
N
N
N
N
R
N
H
O
N
N
N
N
7, 8
H
H
11
9, 10
Scheme 1 Synthesis of compounds 2–10. R: 2, 5, 7, 10 C6H5–; 3, 8, 11 4-Cl-C6H4–; 4, 9 CH3–; 6, 2,6-
(CH3)2-C6H3–. Reagents and conditions: (a) MeOH, RC6H4CNO, reflux, 10–15 min (59–95 %);
(b) MeOH, RC6H4CNS, reflux, 20 min–8 h (55–78 %); (c) POCl3, reflux 24 h (54–73 %); (d) KOH,
reflux, 4 h (61–90 %); (e) MeOH, H2SO4, rt, 15–30 min (65–79 %). Reagents and conditions: (a) MeOH,
RC6H4CNS, reflux, 20 min–8 h (55–78 %); (b) MeOH, RC6H4CNO, reflux, 10–15 min (59–95 %);
(c) POCl3, reflux 24 h (54–73 %); (d) MeOH, H2SO4, rt, 15–30 min (65–79 %); (e) KOH, reflux, 4 h
(61–90 %); (f) MeOH, NH2NH2 H2O, rt, 12 h (70 %)
its chloro analogue 8 were synthesized by heating bisphenylsemicarbazides 2 and 3
under reflux in phosphoryl chloride. Thiadiazole derivatives 9 and 10 were
synthesized from bisthiosemicarbazides 4 and 5 by stirring their mixture in
concentrated sulfuric acid at room temperature for 15 min. The formation of
oxa(thia)diazole ring in compounds 7–10 has been confirmed by the absence of the
proton signals attributable to the CONHNH group in the 1H NMR spectra in
comparison with the spectra for 2–5.
Cyclization of bissemicarbazides 3–6 in basic medium resulted in formation of
5,50-((phenylazanediyl)bis(ethane-2,1-diyl))bis(4-(4-chlorophenyl)-2,4-dihydro-3H-
1,2,4-triazol-3-one) (11), and thio derivatives 12–14 (Scheme 2). The resonances of
hydrogen atoms of the methylene group in the 1H NMR spectra of 11–14 are found
shifted down-field in comparison with the corresponding signals in the spectra of the
starting bissemicarbazides 3–6 due to the significant deshielding effect of the
neighboring triazole ring.
Depending on the pH of the medium, triazolethiones can exist in thione form or
thiole one. The products of triazolethione alkylation and acylation reactions have
been isolated from the reaction mixtures only under basic conditions when thione-
thiol equilibrium of triazolethiones is shifted towards the formation of thioles.
Alkylation reactions of bistriazolethione 13 with haloalkanes, in DMF in the
presence of NaH at room temperature, furnished compounds 15–20 in 50–70 %
yield. Alkylation of 13 with 2-chloroacetamide, chlorophenylmethane or ethyl
123

_
I. Tumosiene et al.
H
N
S
S
H
N
N
N
O
N
H
N
N
N
H
R
R
R
a
N
N
S
H
N
R
N
H
N
H
O
S
N
H
4 - 6
12 - 14
O
N
N
S
S
NH2
N
N
R'
R'
N
H
N
N
N
R
b
R
N
N
R
R
N
H
N
N
N
NH2
S
S
N
N
O
15 - 26
27
Scheme 2 Synthesis of compounds 12–27. R: 5, 13, 15–27 C6H5–; 4, 12 CH3–; 6, 14 2,6-(CH3)2-C6H3–;
R0: 15 CH3CH2–; 16 (CH3)2CH–; 17 (CH3)2CHCH2–; 18 CH3CH2CH2CH2–; 19 (CH3)2CHCH2CH2–; 20
CH3CH2CH2CH2CH2–; 21 NH2COCH2–; 22 C6H5CH2–; 23 C6H5COCH2–; 24 4-ClC6H4COCH2–; 25
4-NO2C6H4COCH2–; 26 C2H5OCOCH2–. Reagents and conditions: (a) KOH, reflux, 4 h (61–90 %);
(b) MeOH, NH2NH2 H2O, rt, 12 h (70 %)
chloroacetate in the presence of KOH and K2CO3 provided bistriazole derivatives 21,
22 and 26, respectively. Bisthiophenyltriazoles 23–25 were synthesized by the
reaction of bistriazolethione 13 with 2-bromoacetophenones in the basic medium
provided by triethylamine. The starting compound 13 is only slightly soluble, whereas
1
the formed S-alkyl derivatives 15–26 dissolve well in common organic solvents. H
NMR spectra of the synthesized S-substituted derivatives 15–26, the resonance
attributable to the NH group proton in the heterocyclic moiety (at approx. 13.71 ppm)
is absent in comparison with the spectrum of bistriazolethione 13. Compound 26 has
been characterized by the presence of additional signals derived from the ester group,
1
which are observed in the H NMR spectrum at 1.19 ppm (-OCH2CH3), and 4.11
(-OCH2CH3) ppm, respectively. Resonance of the NH group proton is shifted down-
field in comparison with the spectrum of the starting compound 13. Carbons of ester
groups resonated at 13.9, and 61.2 ppm, respectively, in the 13C NMR spectrum. The
signal of CO carbon is observed at 153.8 ppm.
Ester 26, upon treatment with hydrazine hydrate, was converted to 2,20-
((((phenylazanediyl)bis(ethane-2,1-diyl))bis(4-phenyl-4H-1,2,4-triazole-5,3-diyl))
bis(sulfanediyl))di(acetohydrazide) (27). In the 1H NMR spectrum for 27, the
signals that originated from the ester functionality in 26 disappeared; instead, new
signals due to the hydrazide structure were recorded at 4.28 and 9.37 ppm,
respectively. Further support for the formation of the hydrazide structure has been
the appearance of strong absorption vibrations indicating the presence of NHNH2 at
3252–3051 cm-1 in the FTIR spectrum.
123

Synthesis and biological activity of 1,3,4-oxa(thia)diazole…
Biology
Evaluation of antioxidative properties
The ability of compounds 2–27 to act as free radical scavengers has been tested by
DPPH (2,2-diphenyl-1-picrylhydrazyl) free radical assay. The presence of the
antioxidant molecule in the reaction mixture with DPPH is indicated by color
change from purple to yellow. Cefazolin, a widely used first-generation
cephalosporin antibiotic, with high antioxidative action, was used as a control. As
seen from the results of the antioxidative activity test (Table 1), the best free radical
scavenging activity has been shown by bistriazolethione 13 containing phenyl
Table 1 Antioxidative and
reducing properties of
compounds 2–27
Compound
DPPH (%)
Absorption
Concentration (mg/ml)
0.5
1
2
13.95
0.39
1.253
0.100
0.208
0.539
1.007
0.074
0.080
0.004
0.049
0.019
0.409
0.019
0.032
0.114
0.101
0.093
0.137
0.084
0.005
0.002
0.033
0.025
0.030
0.190
0.064
0.844
–
0.532
0.187
1.493
1.423
1.128
0.108
0.275
0.058
0.220
0.092
1.077
0.275
0.081
0.171
0.165
0.145
0.190
0.145
0.036
0.017
0.057
0.057
0.067
0.235
0.084
1.165
–
3
4
10.85
27.86
32.17
74.37
52.99
8.68
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
Cefazolin
28.44
22.46
18.26
95.09
19.16
51.45
50.00
52.60
44.80
43.35
48.96
50.62
3.61
79.97
87.34
75.66
2.06
59.02
32.06
123

_
I. Tumosiene et al.
substituent. When this substituent was replaced by the methyl or dimethyl radical,
antioxidative activity decreased 5 times. Activity of S-substituted derivatives of
bistriazolethione 13 depends on the nature of the substituent. The free radical
scavenging activity of compounds containing alkyl groups (15–20) was approx.
50 %, whereas that of 22 containing benzyl group was only 3.61 %. The presence of
phenacyl or 4-substituted phenacyl moiety in the structure of bisthiophenyltriazoles
23, 24, and 25 enhanced the antioxidative activity as well. The positive
antioxidative effect was reinforced by the chloro substituent (24). Replacement of
the ester moiety in a very inactive derivative 26 by the hydrazide moiety (27)
resulted in a 28 times higher antioxidative activity.
Evaluation of reducing properties
Reducing properties of the synthesized compounds 2–27 were evaluated by testing
their power to reduce ferric ion (Fe?3) to ferrous ion (Fe?2). In this method,
reducing agent forms a colored complex with potassium ferricyanide, trichloro
acetic acid and ferric chloride. The higher is the absorption, the stronger reducing
properties of the molecule. The synthesized derivatives are weak reducing agents.
Among them, bisthiosemicarbazides possessing methyl group (4) and phenyl one
(5) were identified as the strongest reducing agents at concentration 1 mg/ml,
whereas diacetamide 21 showed the weakest reducing power (Table 1).
Evaluation of growth regulating properties
The growth regulating activity of the triazolethione 13 and its S-substituted
derivatives 15–20, 22, 26, and 27 was screened on rapeseed cultivar ‘Terra’
(Table 2). The growth regulating properties of S-substituted triazoles depend on the
Table 2 Results of investigation of growth regulating properties of compounds 13, 15–20, 22, 26, and 27
Concentration Control
Compound
13 15
16
17
18
19
20
22
26
27
0.5 mg/l
Average height of seedlings (mm)
27.1 25.3 30.2 26.8 30.1 31.5 31.2 28.2 28.2 30.8 26.8
Average length of roots (mm)
72.9 55.5 61.1 58.7 65.0 60.6 59.2 58.4 51.5 58.9 53.8
Average number of leaves
3.9 3.6
Average height of seedlings (mm)
27.1 24.6 31.8 29.1 31.9 34.5 34.8 32.2 28.3 34.4 29.7
Average length of roots (mm)
72.9 54.5 58.9 55.5 61.4 60.1 55.2 55.4 47.9 57.6 50.5
Average number of leaves
3.9 3.5
3.8
3.8
3.9
3.9
3.9
3.8
3.9
3.8
3.8
1 mg/l
3.9
3.7
3.9
3.8
3.8
3.6
3.7
3.7
4.1
123

Synthesis and biological activity of 1,3,4-oxa(thia)diazole…
nature of the substituent. Bisphenyltriazoles 18 and 19 had the best effect on growth
of the seedlings at both concentrations. Bisisobuthylthiophenyltriazole 17 proved to
be the best stimulator of root growth independent of concentration. However, the
results were worse than those of the control sample. The tested compounds did not
affect significantly the number of the leaves in comparison with the control sample.
Table 3 Minimum inhibitory concentration (MIC, lg/ml) and minimum bactericidal concentration
(MBC, lg/ml) (minimum fungicidal concentration values, MFC for C. albicans) values for the tested
compounds 1–27
Compound Staphylococcus
aureus
Escherichia coli
Pseudomonas
aeruginosa
Candida albicans
MIC
(lg/ml)
MBC
(lg/ml)
MIC
(lg/ml)
MBC
(lg/ml)
MIC
(lg/ml)
MBC
(lg/ml)
MIC
(lg/ml)
MFC
(lg/ml)
1
125
125
250
125
125
125
125
125
62.5
62.5
125
125
62.5
62.5
125
250
250
250
125
31.25
125
125
125
125
125
62.5
125
62.5
250
250
500
250
500
250
250
250
125
125
250
250
125
62.5
125
250
250
250
250
31.25
250
250
125
250
125
125
125
62.5
250
250
250
250
250
250
250
250
250
250
250
250
250
250
500
500
500
500
250
250
250
250
250
250
125
250
125
250
500
500
500
500
500
500
500
250
500
500
500
500
500
500
500
500
500
500
250
250
500
250
250
250
125
250
250
250
500
250
250
250
250
250
250
500
500
500
500
500
500
500
250
125
125
125
125
?
500
500
500
500
500
500
500
500
500
500
500
500
500
500
500
250
125
125
125
?
125
500
125
500
125
500
250
62.5
31.25
125
31.25
62.5
125
125
3.9
250
500
250
500
250
500
500
62.5
31.25
250
31.25
125
250
250
7.3
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
C*
3.9
7.3
3.9
7.3
7.3
15.6
7.3
3.9
3.9
7.3
125
62.5
62.5
62.5
125
7.3
250
62.5
125
62.5
250
7.3
3.9
7.3
3.9
7.3
3.9
7.3
3.9
7.3
3.9
7.3
3.9
7.3
62.5
250
62.5
250
3.9
7.3
15.6
15.6
* Oxytetracycline was used as a control for S. aureus, E. coli, and P. aeruginosa. Nystatin was used as a
control for C. albicans
?, growth of microorganisms
123

_
I. Tumosiene et al.
Evaluation of antimicrobial activity
The antimicrobial activity of the compounds 1–27 was screened by testing their
different concentrations against gram-positive cocci Staphylococcus aureus (ATCC
9144), Gram-negative rods Escherichia coli (ATCC 8739), Pseudomonas aerug-
inosa (NCTC 6750) and fungi Candida albicans (ATCC 10231) by the broth and
spread-plate methods. The minimum inhibition concentration (MIC, lg/ml) and
minimum bactericidal concentration values (MBC, lg/ml) (minimum fungicidal
concentration values, MFC for C. albicans) are listed in the Table 3. A broad-
spectrum antibiotic oxytetracycline was used as a control for S.aureus, E. coli, and
P. aeruginosa. Nystatin was used as a control for C. albicans. As the screening data
for antibacterial activity have shown, a number of the investigated compounds
possess antibacterial properties. Bispentylthiophenyltriazole 20 has shown a very
good bactericidal activity against S. aureus with MIC and MBC values of 31.25 lg/ml,
which are lower than the ones for oxytetracycline. Antimicrobial activity of this
class of compounds depends on the nature of S-substituent. Compounds 9, 10, 13,
14, and 26 inhibit growth of this bacteria strain at 62.5 lg/ml and their MBC values
are 125 (62.5 lg/ml for bisdimethylphenyltriazolethione 14; the same as the one for
oxytetracycline). Bisphenyltriazolesulfaneylacetate 26 has proven to be exception-
ally active against P. aeruginosa with MIC and MBC values of 7.3 lg/ml in
comparison to the values for oxytetracycline of 250 lg/ml. Strain P. aeruginosa
was susceptible to bisphenylthiotriazoles 22, 23, 24, and 27 at MIC value of
62.5 lg/ml. Among tested compounds, just 20 did not suppress growth of this
bacterial strain. Triazoles (15–17 and 19–27) have been tested to possess
exceptional fungicidal activity against fungi C. albicans with MIC value of
3.9 lg/ml and MFC value of 7.3 lg/ml. MIC and MFC values for the precursors of
these derivatives are 125 and 250 lg/ml, respectively.
Conclusions
In summary, a series of 3-((2-carboxyethyl)phenylamino)propanoic acid derivatives
bearing two identical heterocyclic moieties were synthesized. Some of the
synthesized compounds have been proven to possess significant biological
activities. Bistriazolethione 13 has shown excellent antioxidant activity of 95 %.
Just slightly lower free radical scavenging activity has been identified for S-
alkylated triazole derivatives possessing phenacyl fragment (23–25) in their
structure. S-alkylated triazole derivatives 15–27 exhibit significant antimicrobial
activity. Staphylococcus aureus bacterial strain has been tested to be susceptible to
pentylthiophenyltriazole 20 with MIC and MBC values of 31.25 lg/ml, twice lower
than those of oxytetracycline. S-Alkylated triazole derivative containing ester
moiety has exhibited excellent bactericidal activity against Pseudomonas aerugi-
nosa with MBC and MIC values 7.3 lg/ml, 34 times lower than those of
oxytetracycline. S-alkylated triazole derivatives 15–27 have displayed significant
fungicidal activity against Candida albicans with MIC value 3.9 lg/ml.
123

Synthesis and biological activity of 1,3,4-oxa(thia)diazole…
References
1. J. Bostrom, A. Hogner, A. Llinas, E. Wellner, A.T. Plowright, J. Med. Chem. 55, 1817 (2012)
2. W. Xu, S. Yang, P. Bhadury, J. He, M. He, L. Gao, D. Hu, B. Song, Pest. Biochem. Physiol. 10, 6
(2011)
3. Y. Hu, C.-Y. Li, X.-M. Wang, Y.-H. Yang, H.-L. Zhu, Chem. Rev. 114, 5572 (2014)
4. R.A. Rane, S.D. Gutte, N.U. Sahu, Bioorg. Med. Chem. Lett. 22, 6429 (2012)
5. J. Sun, M.-H. Li, S.-S. Qian, F.-J. Guo, X.-F. Dang, X.-M. Wang, Y.-R. Xue, H.-L. Zhu, Bioorg.
Med. Chem. Lett. 23, 2876 (2013)
ˇ
_
_
ˇ
_
ˇ
6. I. Tumosiene, K. Kantminiene, A. Pavilonis, Z. Mazeliene, Z.J. Beresnevicius, Heterocycles 78, 59
(2009)
7. K.I. Bhat, K.S. Sufeera, K.P. Chaitanya, J. Young Pharm. 3, 310 (2011)
8. P. Singh, P.K. Jangra, Der Chem. Sin. 1, 118 (2010)
9. B. Kumar, V. Raj, A. Kumar, V. Singh, Int. J. Curr. Pharm. Res. 4, 9 (2012)
10. Y. Kotaiah, N. Harikrishna, K. Nagaraju, C.V. Rao, Eur. J. Med. Chem. 58, 340 (2012)
11. M. Juszczak, J. Matysiak, M. Szeliga, P. Pozarowski, A. Niewiadomy, J. Albrecht, W. Rzeski,
Bioorg. Med. Chem. Lett. 22, 5466 (2012)
12. M.N. Noolvi, H.M. Patel, S. Kamboj, A. Kaur, V. Mann, Eur. J. Med. Chem. 56, 56 (2012)
´
13. T. Plech, M. Wujec, U. Kosikowska, A. Malm, B. Kapron, Eur. J. Med. Chem. 47, 580 (2012)
14. S.-F. Barbuceanu, G. Saramet, G.L. Almajan, C. Draghici, F. Barbuceanu, G. Bancescu, Eur. J. Med.
Chem. 49, 417 (2012)
15. B. Chandrakantha, A.M. Isloor, P. Shetty, H.K. Fun, G. Hegde, Eur. J. Med. Chem. 71, 316 (2014)
16. A. Cansız, A. Cetin, C. Orek, M. Karatepe, K. Sarac, A. Kus, P. Koparir, Spectrochim. Acta A Mol.
Biomol. Spectrosc. 97, 606 (2012)
17. V. Mathew, J. Keshavayya, V.P. Vaidya, M.H. Moinuddin, Khan. J. Coord. Chem. 61, 2629 (2008)
18. E.S. Il’inykh, D.G. Kim, Chem. Heterocycl. Com. 47, 636 (2011)
ˇ
19. I. Tumosiene, I. Jonuskiene, K. Kantminiene, Z.J. Beresnevicius, Monatsh. Chem. 145, 319 (2014)
20. J. Stefanska, M. Struga, S. Tyski, J. Kossakowski, M. Dobosz, Pol. J. Microbiol. 57, 179 (2008)
ˇ
_
_
_
´
21. R. Nunna, D. Ramachandran, V.B. Modi, K.J. Goswami, Der. Pharma Chemica 4, 639 (2012)
22. G.W. Ware, D.M. Whitacre, An. Introduction to Herbicides (2nd ed.) (Extracted from The Pesticide
Book, 6th ed. (2004), Published by MeisterPro Information Resources, A division of Meister Media
Worldwide, Willoughby, Ohio, USA), http://ipmworld.umn.edu/chapters/whitacreherb.htm. Acces-
sed 30 June 2015
23. K.D. Patel, S.M. Prajapati, S.N. Panchal, H.D. Patel, Synthetic Commun. 44, 1859 (2014)
24. S.C. Holm, B.F. Straub, Org. Prep. Proc. Int. 43, 319 (2011)
_
ˇ
25. I. Tumosiene, Z.J. Beresnevicius, Monatsh. Chem. 140, 1523 (2009)
_
ˇ
_
_
ˇ
26. I. Tumosiene, I. Jonuskiene, K. Kantminiene, Z.J. Beresnevicius, Monatsh. Chem. 143, 1441 (2012)
27. I. Tumosiene, Z.J. Beresnevicius, Chem. Heterocycl. Comp. 43, 1148 (2007)
28. L.N. Madhu, K.N. Suchetha, B.K. Sarojini, DIT 3, 297 (2011)
29. V. Jaishree, N. Ramdas, J. Sachin, B. Ramesh, J. Saudi Chem. Soc. 16, 371 (2012)
30. A.M. Snow, A.E. Ghaly, Am. J. Environ. Sci. 4, 89 (2008)
ˇ
31. K.A. Sazanova, D.I. Bashmakov, A. Brazaityte, C. Bobinas, P. Duchovskis, A.S. Lukatkin, Zem-
_
dirbyste-Agriculture 99, 273 (2012)
32. A. Franklin, J. Acar, F. Anthony, R. Gupta, T. Nicholls, Y. Tamura, S. Thompson, E.J. Thelfall, D.
Vose, M. van Vuuren, D.G. White, H.C. Wegener, M.L. Costarrica, Rev. Sci. Tech. Off. Int. Epiz. 20,
859 (2001)
33. B. Goodson, A. Ehrhardt, S. Ng, J. Nuss, K. Johnson, M. Giedlin, R. Yamamoto, A. Krebber, M.
Ladner, M.B. Giacona, C. Vitt, J. Winter, Antimicrob. Agents Chemother. 43, 1429 (1999)
¨
34. A. Petersen, F.M. Aarestrup, M. Hofshagen, H. Sipila, A. Franklin, E. Gunnarsson, Microb. Drug
Resist. 9, 381 (2003)
123