Thiazole-based chalcones as potent antimicrobial agents. Synthesis and biological evaluation

Article abstract of DOI:10.1016/j.bmc.2011.04.007

As part of ongoing studies in developing new antimicrobials, we report the synthesis of a new class of structurally novel derivatives, that incorporate two known bioactive structures a thiazole and chalcone, to yield a class of compounds with interesting

Full text of DOI:10.1016/j.bmc.2011.04.007

Bioorganic & Medicinal Chemistry 19 (2011) 3135–3140
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Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Thiazole-based chalcones as potent antimicrobial agents. Synthesis
and biological evaluation
K. Liaras a, A. Geronikaki a, , J. Glamoclija , A. Ciric , M. Sokovic
⇑
b
b
b
´
ˇ
´
´
a Aristotle University, School of Pharmacy, Department of Pharmaceutical Chemistry of Aristotle University of Thessaloniki, Thessaloniki 54124, Greece
b Institute for biological research ‘S.Stankovi´c’, Mycological laboratory, University of Belgrade, 11000, Serbia
a r t i c l e i n f o
a b s t r a c t
Article history:
As part of ongoing studies in developing new antimicrobials, we report the synthesis of a new class of
structurally novel derivatives, that incorporate two known bioactive structures a thiazole and chalcone,
to yield a class of compounds with interesting antimicrobial properties.
Evaluation of antibacterial activity showed that almost all the compounds exhibited greater activity
than reference drugs and thus could be promising novel drug candidates.
Received 3 January 2011
Revised 29 March 2011
Accepted 1 April 2011
Available online 7 April 2011
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
Thiazole
Chalcones
Antibacterial
Antifungal
1. Introduction
Gram positive and Gram negative bacteria and also against a ser-
ies of fungi.
Despite the rapid progress of science, the treatment of infec-
tious diseases still remains a serious problem of concern to the sci-
entific community, mainly because of the wide range of factors
leading to the emergence of these diseases and also the increased
number of pathogenic microorganisms with resistance to multiple
drugs, including the Gram positive bacteria.1–5
To limit the emerging resistance of microorganisms requires the
careful use of existing antimicrobial drugs. However, there is a
need for the design of novel antimicrobial agents, particularly for
the treatment of the infections of hospitalized patients and protec-
tion of immunosuppressed or HIV-infected patients.
2. Results and discussion
2.1. Chemistry
The synthesis of the target chalcones 1–10 was accomplished by
a Claisen–Schmidt condensation of 1-(4-methyl-2-(methylamino)
thiazol-5-yl)ethanone with various aromatic aldehydes, in metha-
nol, and 10% aq NaOH,24 as indicated to Scheme 1. The reactions
R
A potential approach is the design of innovative drugs, with dif-
ferent mechanism of action, in effort to avoid cross resistance to
existing therapeuticals.6 Recently, linezolid,7–9 a synthetic mole-
cule containing oxazolidinone moiety has been approved for clini-
cal use. It is effective against Gram positive bacteria while
appearing to have a unique mechanism of action.
CH3
N
H3C
C
O
N
S
H
Our design supported from our previous findings10–15 addressed
the synthesis of a new class of structurally novel derivatives, that
incorporate two known bioactive structures a thiazole16–26 and a
chalcone,27–29 to yield a class of compounds with interesting anti-
microbial properties
1.
2.
3.
4.
5.
Η
6. 2-Cl
4-NO2
3-NO2
4-Cl
7. 4-OMe
8. 2-OMe
9. 2,6-diCl
10. 2,4-diCl
Ten new thiazole-based chalcones (Fig. 1) were synthesized
and tested for their in vitro antimicrobial properties against
3-Cl
⇑
Corresponding author. Tel.: +30 2310 997616; fax: +30 2310 998559.
E-mail address: geronik@pharm.auth.gr (A. Geronikaki).
Figure 1. Structure of the synthesized compounds.
0968-0896/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2011.04.007

3136
K. Liaras et al. / Bioorg. Med. Chem. 19 (2011) 3135–3140
O
O
NH2
N
-HCl
-H2O
+
N
S
N
S
H
H
O
Cl
R
CH3
N
R
N
NaOH 10%
+
OHC
N
H3C
S
H
C
N
O
S
H
O
Scheme 1. Synthesis of title compounds.
Table 1
positive and Gram negative bacteria, and molds and the minimal
inhibitory concentrations that inhibited the growth of the tested
microorganisms (MIC) and minimal bactericidal/fungicidal con-
centration were determined. The results of antimicrobial testing
against a panel of selected Gram positive and Gram negative bac-
teria are reported in Table 2, along with those of reference drugs
Ampicillin and Streptomycin.2
Calculated lipophilicity of synthesized compounds expressed as C log P, A Log Ps ja
A Log P
i
R
CH3
N
All the compounds tested have activity against all the bacteria
tested roughly comparable to that of Ampicillin, and rather less
than of Streptomycin and minimal inhibitory concentration (MIC)
H3C
C
O
N
S
H
is at range of 7.64–45.87
concentration (MBC) is 30.58–68.37
to this generalization are the compound 6 possesses threefold
l
mol/ml ꢁ 10ꢀ2 and minimal bactericidal
F/F
C log Pa
C log Pb
A Log Psc
A Log Pd
l
mol/ml ꢁ 10ꢀ2. Exceptions
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
3.43
3.17
3.17
4.14
4.14
4.14
3.57
3.57
4.29
3.93
3.35
3.35
4.73
4.85
4.85
2.97
2.91
2.91
3.57
3.57
3.57
3.11
3.11
3.74
3.48
2.81
2.81
4.18
4.18
4.18
3.41
3.02
3.15
3.87
3.80
3.80
3.46
3.46
3.96
3.71
3.49
3.43
4.05
4.04
3.93
3.08
2.98
2.98
3.75
3.75
3.75
3.29
3.29
3.83
3.57
3.07
3.07
4.41
4.41
4.41
better MIC against Enterococcus Faecalis than does Ampicillin,
and the compounds
4 and 7 possess threefold better MIC
against Micrococcus flavus than does Ampicillin. Probably these
two bacteria are more sensitive to title compounds than to
Ampicillin.
The lowest activity was observed for compound 4 against Staph-
ylococcus aureus with MIC 51.28 ꢁ 10ꢀ2 and MBC 68.37
lmol/
ml ꢁ 10ꢀ2
.
It was found that the most sensitive bacteria to compounds
tested was Gram negative bacteria E. faecalis, while, Gram-positive
Listeria monocytogenes, was the most resistant one.
It is noteworthy, that all tested compounds exhibited activity
equal or better than commercial antibiotic Ampicillin but they
were less active compared to Streptomycin. DMSO in used concen-
tration did not show any effect on bacteria.
a
Calculated by ChemDraw Ultra 10.0.
Calculated by online program of ChemAxon.
Calculated by online program ALOGPS 2.1.
b
c,d
The results of antifungal activity are presented in Table 3. All the
compounds testedpossess modestactivityagainst all thefungalspe-
cies tested; but that either ketoconazole or bifonazole, or both, are
more active than any of the chalcones tested. The antifungal proper-
ties of the chalcones are rather broad spectrum: all the activities
determined were within a threefold concentration range. Notewor-
thy results are that: (i) Compounds 2, 4, 5, 6, 8, 9, and 10 are all only
1.5- to 2-fold less active against Fusarium sporotrichoides compared
to bifonazole, the standard possessing the better activity. (ii) All 10
novel compounds are only twofold less active against Trichoderma
viride than compared to ketoconazole, the standard possessing the
better activity. (iii) Additionally compound 4 is only 3.5-fold less ac-
tive against Aspergillus niger than compared to bifonazole; com-
pound 8 is only fourfold less active against Aspergillus ochraceus
than compared to bifonazole.
proceededsmoothlyandingood yields(32–84%). Allthecompounds
formed were recrystallized from dioxane or ethanol.
All new structures of compounds 1–10 were satisfactorily con-
firmed by IR, 1H NMR and elemental analysis. IR spectra showed
absorptions at 1650–1720 cmꢀ1 (C@O), and sharp bands at
3200 cmꢀ1 (NH). In the 1H NMR spectra chalcones showed peaks
in the region of d 2.54–2.78 (CH3), 2.88–2.98 (N–CH3), 7.14–7.26
(CO–CH) and 7.53–7.58 (Ar-CH).
The C –Cb double bond in the enone moiety of chalcones can
a
potentially adopt either a Z or an T configuration. The 1H NMR
spectrum of each compounds exhibited CH@CH protons around
7.14–7.63 ppm, with J >15, would suggest that the compounds
were produced with an (E) configuration.30
Theoretical calculations of lipophilicity using the method of
additivity to generate C log P values, was performed (Table 1).31
Among compounds tested the best antifungal activity was
expressed by chalcone 6 (MIC 15.29
Aspergillus versicolor, N. viride and F. sporotrichoides, as well as
l
mol/ml ꢁ 10ꢀ2) against
2.2. Biological evaluation
compound
9 against N. viride, Penicillium ochrochloron and
2.2.1. Antimicrobial activity
The synthesized thiazole-based chalcones were then assayed
in vitro for their antibacterial and antifungal activity against Gram
F. sporotrichoides., even though these compounds are less potent
than reference drugs (ketoconazole and bifonazole). Quite good
antifungal activity was observed for compounds 7 and 8 while

K. Liaras et al. / Bioorg. Med. Chem. 19 (2011) 3135–3140
3137
Table 2
Antibacterial activity of substituted/non substituted (E)-1-[4-methyl-2-(methylamino)thiazol-5-yl)-3-phenylprop-2-en-1-ones (MIC and VDC,
l
mol/ml ꢁ 10ꢀ2
)
R
CH3
N
H3C
C
O
N
S
H
Compounds
S.a.
B.c.
M.f.
L.m.
Ps. aer.
S. typhi
E.coli
En.f.
1
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
MIC
MBC
38.75
58.13
33.00
49.50
33.00
49.50
51.28
68.37
34.18
51.28
30.58
45.87
29.05
34.18
34.72
34.72
30.58
45.87
30.58
45.87
24.79
37.19
1.71
38.75
58.13
33.00
49.50
33.00
33.00
34.18
51.28
34.18
51.28
22.93
30.58
29.05
34.18
34.72
34.72
25.99
30.58
22.93
30.58
24.79
37.19
0.20
19.37
38.75
33.00
49.50
33.00
49.50
8.54
34.18
25.64
34.18
22.93
25.99
8.54
17.09
26.04
29.51
22.93
30.58
30.58
45.87
24.79
37.19
0.20
38.75
58.13
33.00
49.50
33.00
49.50
34.18
51.28
34.18
51.28
30.58
45.87
34.18
51.28
34.72
52.08
30.58
45.87
30.58
45.87
24.79
37.19
1.71
29.06
38.75
33.00
49.50
28.05
33.00
25.64
34.18
34.18
51.28
25.99
30.58
34.18
51.28
29.51
34.72
22.93
30.58
30.58
30.58
24.79
37.19
0.20
32.94
38.75
24.75
33.00
28.05
33.00
29.05
34.18
29.05
34.18
30.58
45.87
34.18
51.28
34.72
52.08
30.58
30.58
22.93
30.58
24.79
37.19
0.20
32.94
38.75
24.75
33.00
24.75
33.00
29.05
34.18
29.05
34.18
22.93
30.58
34.18
51.28
26.04
34.72
22.93
30.58
30.58
45.87
24.79
37.19
0.86
19.37
38.75
16.50
33.00
33.00
33.00
17.10
34.18
8.54
2
3
4
5
34.18
7.64
6
15.29
17.09
17.09
26.04
34.72
15.29
30.58
30.58
45.87
24.79
37.19
0.43
7
8
9
10
Ampicillin
Streptomycin
4.29
0.43
0.43
4.30
0.43
0.43
1.71
0.86
S.a.—Staphylococcus aureus (ATCC 6538); B.c.—Bacillus cereus (clinical isolate); M.f.—Micrococcus flavus (ATCC 10240); L.m.—Listeria monocytogenes (NCTC 7973); Ps. aer.—
Pseudomonas aeruginosa (ATCC 27853); S. typhi—Salmonella typhimurium (ATCC 13311); E.coli—Escherichia coli (ATCC 35210); En.f.—Enterococcus faecalis (human isolate).
Table 3
Antifungal activity of substituted/non substituted (E)-1-[4-methyl-2-(methylamino)thiazol-5-yl)-3-phenylprop-2-en-1-ones (MIC and MFC in
l
mol/ml ꢁ 10ꢀ2
)
Compounds
A.o.
A.v.
19.38
A.n.
A.f.
T.v.
19.38
P.o.
19.38
P.f.
F.s.
1
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
38.80
58.10
25.00
33.00
33.00
49.60
34.00
51.30
34.00
51.30
30.58
45.87
34.00
51.30
17.00
34.00
45.87
61.16
30.58
61.16
4.70
38.80
77.52
33.00
82.50
33.00
66.00
17.00
34.00
34.00
85.40
30.58
61.16
34.00
68.30
34.00
87.00
30.58
61.16
45.87
61.16
28.25
47.00
5.03
38.80
58.10
33.00
49.60
33.00
49.60
17.00
34.00
34.00
51.30
30.58
61.16
17.00
51.30
34.00
52.00
30.58
30.58
15.29
61.16
4.70
38.80
58.10
33.00
49.60
33.00
49.60
17.00
34.00
34.00
51.30
30.58
61.16
34.00
51.30
34.00
69.00
30.58
45.87
15.29
30.58
0.94
38.80
38.80
16.50
66.00
33.00
66.00
17.00
17.00
17.00
68.30
15.29
45.87
34.00
51.30
17.00
34.00
15.29
45.87
15.29
30.58
9.42
58.10
33.00
49.60
33.00
49.60
17.00
34.00
34.00
51.30
15.29
45.87
17.00
51.30
34.00
52.00
30.58
45.87
15.29
61.16
1.88
38.80
16.50
33.00
16.50
33.00
17.00
34.00
17.00
34.00
15.29
30.58
17.00
51.30
17.00
34.00
15.29
30.58
15.29
30.58
9.42
77.52
33.00
49.60
33.00
49.60
34.00
51.30
34.00
51.30
30.58
45.87
34.00
51.30
34.00
52.00
15.29
45.87
30.58
61.16
1.88
2
3
4
5
6
7
8
9
10
Ketoconazole
Bifonazole
9.42
40.30
80.60
4.70
2.51
161.16
9.42
0.32
6.12
18.80
80.60
161.16
4.70
40.30
161.16
1.88
161.16
241.74
18.80
8.06
80.60
20.14
A.o.—Aspergillus ochraceus (ATCC 12066); A.v.—Aspergillus versicolor (ATCC 11730); A.n.—Aspergillus niger (ATCC 6275); A.f.—Aspergillus fumigatus (plant isolate); T.v.—Trich-
oderma viride (IAM 5061); P.o.—Penicillium ochrochloron (ATCC 9112); P.f.—Penicillium funiculosum (ATCC 36839); F.s.—Fusarium sporotrichoides (IMT 496).
the lowest activity was observed for chalcone 1, with MIC
As regards the relationships between the structure of the het-
erocyclic scaffold and the detected antibacterial properties, clearly,
the introduction of the bulky electron-withdrawing nitro and
19.38–38.80
l
mol/ml ꢁ 10ꢀ2. DMSO in used concentration did
not show any effect on fungi.

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K. Liaras et al. / Bioorg. Med. Chem. 19 (2011) 3135–3140
chloro groups enhances the antibacterial properties. It was ob-
served that the introduction of methoxy group is favored too for
antibacterial activity (compounds 7 and 8) with respect to unsub-
stituted derivative 1. In generally, compounds 4, 6, 7, and 8 stand
out as the compounds possessing the more noteworthy properties,
which are either mono-chloro or mono-methoxy substituted.
It seems also interesting to point out that, among the isomeric
chloro substituted compounds 4–6, as well as among the nitro
substituted 2–3, the position of the substituent exerts, in general,
a certain effect, even if not univocal, on the activity against all
the microorganisms. Conversely, among the nitro substituted 2,
3, the meta—derivative (compound 3) are endowed with the higher
activity, whereas, for the chloro substituted compounds 4–6 it is
evident the greater efficacy of the ortho derivative 6 with respect
to m- and p-derivatives.
Solvents, unless otherwise specified were of analytical reagent
grade or of the highest quality commercially available. Synthetic
starting materials, reagents and solvents were purchased from Al-
drich Chemie (Steinheimm, Germany).
4.1. Synthesis of 1-(4-methyl-2-(methylamino)thiazol-5-
yl)ethanone
1-Methylthiourea (1.8 g, 0.02 mol) was dissolved in 50 ml of
acetone. 3-Chloroacetylacetone (2.26 ml, 0.02 mol), diluted in ace-
tone (5 ml) was added dropwise and mixture was refluxed for
1.5 h. The solid product was filtered and recrystallized from
ethanol.32
4.2. General procedure for the synthesis of (E)-1-(4-methyl-2-
The majority of compounds showed the worst activity against A.
ochraceus followed by P. ochrochloron, while T. viride, followed by
A. versicolor and F. sporotrichoides, are the most sensitive species.
The last two species are in general more sensitive than others used
in experiment.11,14
(methylamino)thiazol-5-yl)-3-phenylprop-2-en-1-ones
1-(4-Methyl-2-(methylamino)thiazol-5-yl)ethanone (1 mol) in
methanol (4.0–4.1 l), was added dropwise to a cooled solution of
corresponding aromatic aldehydes (1 mol) in 10% NaOH (600–
650 ml). The solution was maintained at 0 °C for 1.5 h and then
was allowed to stir at room temperature. After some time (5–
12 h) solid started separating out. The solid was filtered under vac-
uum and recrystallized from dioxane or ethanol to give the title
chalcones. The yield range after recrystallization of chalcones
was 32–84%.
No correlation of antimicrobial activity, and lipophilicity, calcu-
lated as C log P, was observed.
3. Conclusion
The newly synthesized thiazole-based chalcones 1–10 exhibit a
remarkable inhibition of the growth of a wide spectrum of Gram
positive bacteria, Gram negative bacteria and fungi. The most sen-
sitive bacterial species on compounds tested is Gram negative bac-
teria E. faecalis, while, Gram-positive L. monocytogenes, is the most
resistant one. As far as the fungi are concerned, the tested com-
pounds posses modest activity against all the fungal species tested
being less active than ketoconazole and bifonazole. The majority of
compounds showed the worst activity against Aspergillus ochraceus
followed by Penicillium ochrochloron, while T. viride, followed by A.
versicolor and F. sporotrichoides, are the most sensitive species. The
last two species are in general more sensitive than others used in
experiment.11,14
It should be noticed that some compounds tested exhibited bet-
ter activity than commercial antimicrobial agents used as refer-
ence drugs while there were less active than ketoconazole and
bifonazole.
The outstanding properties of this new class of antibacterial
substances deserve further investigation in order to clarify the
mode of action at molecular level, responsible for the activity
observed.
4.2.1. (E)-1-(4-Methyl-2-(methylamino)thiazol-5-yl)-3-phenyl
prop-2-en-1-one (1)
Yield: 32%, mp: 186–187 °C. IR (Nujol) 3250, 1625 (C@C), 1720
;
(C@O), 1600 (C–H arom) cmꢀ1 1H NMR (d ppm, DMSO-d6,
300 MHz): 2.54 (s, 3H, thiazole 40-CH3), 2.88 (s, 3H, N–CH3), 7.14
(d, J = 15.6 Hz, 1H, CO–CH), 7.34–7.59 (m, 6H, phenyl 20 and 60, phe-
nyl 30 and 50, phenyl 40, Ph-CH), 8.31 (s, 1H, thiazole 20-NH). Anal.
Calcd for C14H14N2OS (MW 258): C, 65.09; H, 5.46; N, 10.84. Found:
C, 65.29; H, 5.36; N, 10.90.
4.2.2. (E)-1-(4-Methyl-2-(methylamino)thiazol-5-yl)-3-(4-
nitrophenyl)prop-2-en-1-one (2)
Yield: 58%, mp: 248–249 °C. IR (Nujol) 3390, 3200 (NH), 1722
(C@O), 1653 (CH@CH), 1602 (C–H arom), 1525 (assym. N@O),
1340 (NO2) cmꢀ1 1H NMR (d ppm, DMSO-d6, 300 MHz): 2.58 (s,
;
3H, thiazole 40-CH3), 2.89 (s, 3H, N–CH3), 7.51 (d, J = 15.6 Hz, 1H,
CO–CH), 7.63 (d, J = 15.6 Hz, 1H, Ar-CH), 8.04 (d, J = 9 Hz, 2H, Ar
30 and 50), 8.26 (d, J = 8.7 Hz, 2H. Ar 20 and 60), 8.65 (s, 1H, thiazole
20-NH). Anal. Calcd for C14H13N3O3S (MW 303): C, 55.43; H, 4.32; N,
13.85. Found: C, 55.35; H, 4.20; N, 13.65.
4. Experimental part
4.2.3. (E)-1-(4-Methyl-2-(methylamino)thiazol-5-yl)-3-(3-
nitrophenyl)prop-2-en-1-one (3)
Melting points (°C) were determined with a MELTEMP II capil-
lary apparatus (LAB Devices, Holliston, MA, USA) without correc-
tion. Elemental analyses were performed on a Perkin–Elmer 2400
CHN elemental analyzer and all compounds synthesized were
within a 0.4% of theoretical values. IR spectra were recorded, in Nu-
jol, on a Perkin Elmer Spectrum BX and a DR-8001 Shimadzu. Wave
Yield: 47%, mp: 246–247 °C. IR (Nujol) 3390, 3250 (NH), 1680
(C@O),1652 (CH@CH), 1602, 1525 (assym. N@O), 1350 (NO2)
cmꢀ1 1H NMR (d ppm, DMSO-d6, 300 MHz): 2.49 (s, 3H, thiazole
;
40-CH3), 2.79 (s, 3H, N–CH3), 7.42–7.63 (m, 3H, alkene, Ar 50),
8.15 (dd, 2H, Ar 60, Ar 40), 8.48 (s, 1H, Ar 20). MS: (m/z): 303 (M+,
5%), 289 (4%), 273 (2%), 181 (6%), 171 (7%), 170 (58%), 155
(100%), 151 (25%), 150 (23%), 127 (11%), 114 (10%), 105 (12%), 95
(9%), 86 (30%), 81 (24%), 77 (19%), 69 (42%), 57 (10%), 55 (11%),
51 (14%), 45 (11%), 43 (19%), 42 (10%), 41 (15%). Anal. Calcd for
numbers in the IR spectra are given in cmꢀ1 1H- NMR spectra of
.
the newly synthesized compounds, in DMSO-d6 solutions, were re-
corded on a Bruker AC 300 instrument (Bruker, Karlsruhe, Ger-
many) at 298 K. Chemical shifts are reported as d (ppm) relative
to TMS as internal standard. Coupling constants J are expressed
in Hertz (Center of Instrumental Analysis of the University of
Thessaloniki).
The reactions were monitored by TLC on F254 silica-gel
precoated sheets (Merck, Darmstadt, Germany) and the purified
compounds each showed a single spot.
C14H13N3O3S (MW 303): C, 55.43; H, 4.32; N, 13.85. Found: C,
55.38; H, 4.22; N, 13.75.
4.2.4. (E)-3-(4-Chlorophenyl)-1-(4-methyl-2-(methylamino)
thiazol-5-yl)prop-2-en-1-one (4)
Yield: 60%, mp: 191–193 °C. IR (Nujol) 3200 (NH), 1680 (C@O),
1650 (CH@CH), 1602, 1100 (Cl) cmꢀ1 1H NMR (d ppm, DMSO-d6,
;

K. Liaras et al. / Bioorg. Med. Chem. 19 (2011) 3135–3140
3139
300 MHz): 2.50 (s, 3H, thiazole 40-CH3), 2.82 (s, 3H, N–CH3), 7.27
4.3. Biological evaluation
4.3.1. Antifungal activity
(d, J = 15.6 Hz, 1H, CO–CH), 7.42–7.50 (m, 3H, Ar 30 and 50, Ar-
CH), 7.74 (d, J = 8.4 Hz, 2H, Ar 20 and 60). Anal. Calcd for
C
14H13ClN2OS (MW 292.5): C, 57.43; H, 4.48; Cl, 12.11; N, 9.57.
For the antifungal bioassays, eight fungi were used: Aspergillus
ochraceus (ATCC 12066), Aspergillus fumigatus (plant isolate),
Aspergillus niger (ATCC 6275), A. versicolor (ATCC 11730), Penicil-
lium funiculosum (ATCC 36839), Penicillium ochrochloron (ATCC
9112), Trichoderma viride (IAM 5061) and F. sporotrichoides
(IMT 496). The organisms were obtained from the Mycological Lab-
oratory, Department of Plant Physiology, Institute for Biological Re-
Found: C, 57.40; H, 4.30; N, 9.68.
4.2.5. (E)-3-(3-Chlorophenyl)-1-(4-methyl-2-(methylamino)
thiazol-5-yl)prop-2-en-1-one (5)
Yield: 61%, mp: 209–212 °C. IR (Nujol) 3200 (NH), 1685 (C@O),
1651 (CH@CH), 1602, 1075 (Cl) cmꢀ1 1H NMR (d ppm, DMSO-d6,
;
300 MHz): 2.48 (s, 3H, thiazole 40-CH3), 2.80 (s, 3H, N–CH3), 7.32–
7.46 (m, 4H, CO–CH, Ar 20, Ar 50, Ar 40), 7.72 (dd, 2H, Ar 60, Ar-CH),
8.52 (s, 1H, NH). Anal. Calcd for C14H13ClN2OS (MW 292.5): C,
57.43; H, 4.48; Cl, 12.11; N, 9.57. Found: C, 57.44; H, 4.40; N, 9.60.
search ‘Siniša Stankovic’, Belgrade, Serbia.
´
The micromycetes were maintained on malt agar and the cul-
tures stored at 4 °C and sub-cultured once a month.33 In order to
investigate the antifungal activity of the compounds, a modified
microdilution technique was used.34–36 The fungal spores were
washed from the surface of agar plates with sterile 0.85% saline
containing 0.1% Tween 80 (v/v). The spore suspension was adjusted
with sterile saline to a concentration of approximately 1.0 ꢁ 105 in
4.2.6. (E)-3-(2-Chlorophenyl)-1-(4-methyl-2-(methylamino)
thiazol-5-yl)prop-2-en-1-one (6)
Yield: 32%, mp: 185–187 °C. IR (Nujol) 3200 (NH), 1680 (C@O),
1650 (CH@CH), 1602, 1025 (Cl) cmꢀ1
;
1H NMR (d ppm, DMSO-d6,
a final volume of 100 ll per well. The inocula were stored at 4 °C
300 MHz): 2.50 (s, 3H, thiazole 40-CH3), 2.81 (s, 3H, N–CH3), 7.3
(d, J = 15.3 Hz, 1H, CO–CH), 7.36 (dd, 2H, Ar 40, Ar 50), 7.49 (d,
J = 9.3 Hz, 1H, Ar 60), 7.76 (d, J = 15.6 Hz, 1H, Ar-CH), 7.93 (d,
J = 9.3 Hz, 1H, Ar 30). MS: (m/z): 292 (M+, 100%), 291 (28%), 257
(43%), 236 (9%), 201 (9%), 182 (8%), 181 (73%), 167 (13%), 165
(9%), 155 (33%), 137 (13%), 128 (9%), 102 (14%), 101 (22%), 86
(15%), 81 (14%), 75 (9%), 69 (25%), 57 (9%). Anal. Calcd for
for further use. Dilutions of the inocula were cultured on solid malt
agar to verify the absence of contamination and to check the valid-
ity of the inoculum.
Minimum inhibitory concentration (MIC) determinations were
performed by a serial dilution technique using 96-well microtiter
plates. The compounds investigated were dissolved in 5% DMSO
solution containing 0.1% Tween 80 (v/v) (1 mg/ml) and added in
broth Malt medium with inoculum. The microplates were incu-
bated at Rotary shaker (160 rpm) for 72 h at 28 °C. The lowest con-
centrations without visible growth (at the binocular microscope)
were defined as MICs.
C14H13ClN2OS (MW 292.5): C, 57.43; H, 4.48; Cl, 12.11; N, 9.57.
Found: C, 57.41; H, 4.45; N, 9.55.
4.2.7. (E)-3-(4-Methoxyphenyl)-1-(4-methyl-2-(methylamino)
thiazol-5-yl)prop-2-en-1-one (7)
The fungicidal concentrations (MFCs) were determined by serial
Yield: 29%, mp: 219–223 °C. IR (Nujol) 3200 (NH), 1684 (C@O),
subcultivation of a 2
and inoculated for 72 h, into microtiter plates containing 100 l
broth per well and further incubation 72 h at 28 °C. The lowest
concentration with no visible growth was defined as MFC indicat-
ing 99.5% killing of the original inoculum. DMSO was used as a neg-
ative control, commercial fungicides, bifonazole (Srbolek, Belgrade,
Serbia) and ketoconazole (Zorkapharma, Šabac, Serbia), were used
l
l of tested compounds dissolved in medium
l of
1654 (CH@CH), 1604, 1584, 1250 C-O–C) cmꢀ1
;
1H NMR (d ppm,
DMSO-d6, 300 MHz): 2.61 (s, 3H, thiazole 40-CH3), 2.93 (s, 3H, N–
CH3), 3.87 (s, 3H, OCH3), 7.05 (d, J = 8.7 Hz, 2H, Ar 30 and 50), 7.22
(d, J = 15.3 Hz, 1H, CO–CH), 7.58 (d, J = 15.6 Hz, 1H, Ar-CH), 7.77
(d, J = 8.7 Hz, 2H, Ar 20 and 60). Anal. Calcd for C15H16N2O2S (MW
288): C, 62.48; H,5.50; N, 9.71. Found: C,62.45; H, 5.52; N, 9.71.
as positive controls (1–3000
All experiments were performed in duplicate and repeated
three times.
lg/ml).
4.2.8. (E)-3-(2-Methoxyphenyl)-1-(4-methyl-2-
(methylamino)thiazol-5-yl)prop-2-en-1-one (8)
Yield: 36%, mp: 213–215 °C. IR (Nujol) 3200 (NH), 1670 (C@O),
1643 (CH@CH), 1607, 1247 (C-O–C) cmꢀ1; 1H NMR (d ppm, DMSO-
d6, 300 MHz): 2.54 (s, 3H, thiazole 40-CH3), 2.86 (s, 3H, N–CH3),
3.88 (s, 3H, OCH3), 7.05 (m, 2H, CO–CH, Ar 30), 7.41 (dd, 2H, Ar 50,
Ar 40), 7.75 (dd, 2H, Ar 60, Ar-CH). Anal. Calcd for C15H16N2O2S
(MW 288): C, 62.48; H,5.50; N, 9.71. Found: C,62.47; H, 5.53; N, 9.70.
4.3.2. Antibacterial activity
The following Gram-negative bacteria were used: Escherichia
coli (ATCC 35210), Pseudomonas aeruginosa (ATCC 27853), Salmo-
nella typhimurium (ATCC 13311), E. faecalis (human isolate) and
the following Gram-positive bacteria: Bacillus cereus (clinical
isolate), M. flavus (ATCC 10240), L. monocytogenes (NCTC 7973),
and Staphylococcus aureus (ATCC 6538). The organisms were
obtained from the Mycological Laboratory, Department of Plant
4.2.9. (E)-3-(2,6-Dichlorophenyl)-1-(4-methyl-2-(methylamino)
thiazol-5-yl)prop-2-en-1-one (9)
´
Yield: 84%, mp: 195–197 °C. IR (Nujol) 3200 (NH), 1690 (C@O),
Physiology, Institute for Biological Research ‘Siniša Stankovic’,
Belgrade, Serbia.
1650 (CH@CH), 1607 cmꢀ1
;
1H NMR (d ppm, DMSO-d6, 80 MHz):
2.50 (s, 3H, thiazole 40-CH3), 2.91 (s, 3H, N–CH3), 7.15–7.8 (m,
5H, Ar, alkene). Anal. Calcd for C14H12Cl2N2OS (MW 327): C,
51.39; H, 3.70; Cl, 21.67; N, 8.56; O, 4.89; S, 9.80. Found: C,
51.40; H, 3.73; N, 8.58.
The antibacterial assay was carried out by a microdilution
method34,35 in order to determine the antibacterial activity of com-
pounds tested against the human pathogenic bacteria.
The bacterial suspensions were adjusted with sterile saline to a
concentration of 1.0 ꢁ 105 CFU/ml. The inocula were prepared dai-
ly and stored at +4 °C until use. Dilutions of the inocula were cul-
tured on solid medium to verify the absence of contamination and
to check the validity of the inoculum.
4.2.10. (E)-3-(2,4-Dichlorophenyl)-1-(4-methyl-2-
(methylamino)thiazol-5-yl)prop-2-en-1-one (10)
Yield: 76%, mp: 239–240 °C. IR (Nujol) 3200 (NH), 1680 (C@O),
1650 (CH@CH), 1602, 1100 (Cl) cmꢀ1
;
1H NMR (d ppm, DMSO-d6,
All experiments were performed in duplicate and repeated
three times.
300 MHz): 2.56 (s, 3H, thiazole 40-CH3), 2.88 (s, 3H, N–CH3), 7.39
(d, J = 15.3 Hz, 1H, CO–CH), 7.51 (d, J = 8.4 Hz, 1H, Ar 50), 7.74–
7.78 (m, 2H, Ar 30, Ar-CH), 8.05 (d, J = 8.3 Hz, 1H, Ar 60), 8.64 (s,
1H, NH). Anal. Calcd for C14H12Cl2N2OS (MW 327): C, 51.39; H,
3.70; Cl, 21.67; N, 8.56. Found: C, 51.43; H, 3.69; N, 8.55.
4.3.3. Microdilution test
The minimum inhibitory and bactericidal concentrations (MICs
and MBCs) were determined using 96-well microtiter plates. The

3140
K. Liaras et al. / Bioorg. Med. Chem. 19 (2011) 3135–3140
13. Argyropoulou, I.; Geronikaki, A.; Vicini, P.; Zani, F. ARKIVOC 2009, vi.
Commemorative Issue in Honor of Professor Henk van der Plas on the
occasion of his 80th anniversary..
bacterial suspension was adjusted with sterile saline to
a
concentration of 1.0 ꢁ 105 CFU/ml. Compounds to be investigated
were dissolved in 5% DMSO solution containing 0.1% Tween 80
14. Camoutsis, C.; Geronikaki, A.; Ciric, A.; Sokovic´, M.; Zoumpoulakis, P.; Zervou,
M. Chem. Pharm. Bull. 2010, 58, 160.
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(v/v) (1 mg/ml) and added in broth LB medium (100 ll) with bac-
´
terial inoculum (1.0 ꢁ 104 CFU per well) to achieve the wanted
concentrations. The microplates were incubated at Rotary shaker
(160 rpm) for 24 h at 48 °C. The lowest concentrations without vis-
ible growth (at the binocular microscope) were defined as concen-
trations that completely inhibited bacterial growth (MICs). The
ˇ
J.; Geronikaki, A. Bioorg. Med. Chem. 2010, 18, 426.
16. Geronikaki, A.; Dearden, J.; Filimonov, D.; Galaeva, I.; Garibova, T. L.;
Gloriozova, T.; Krajneva, V.; Lagunin, A.; Macaev, F. Z.; Molodavkin, G.;
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Krajneva, V.; Lagunin, A.; Macaev, F.; Molodavkin, G.; Poroikov, V.; Pogrebnoi,
S.; Saloutin, V.; Stepanchikova, A.; Stingaci, E.; Tkach, N.; Vlad, L.; Voronina, T.
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MBCs were determined by serial sub-cultivation of 2
ll into micro-
titer plates containing 100 l of broth per well and further incuba-
l
tion for 24 h. The lowest concentration with no visible growth was
defined as the MBC, indicating 99.5% killing of the original
inoculum. The optical density of each well was measured at a
wavelength of 655 nm by Microplate manager 4.0 (Bio-Rad Labora-
tories) and compared with a blank and the positive control. Strep-
tomycin (Sigma P 7794) and Ampicillin (Panfarma, Belgrade,
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was used as a negative control.
18. Geronikaki, A.; Eleftheriou, P.; Alam, Intekhab.; Saxena, A. K. J. Med. Chem.
2008, 51, 5221.
19. Kouatly, O.; Geronikaki, A.; Kamoutsis, C.; Hadjipavlou-Litina, D.; Eleftheriou, P.
Eur. J. Med. Chem. 2009, 44, 1198.
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N.; Modarresi, H.; Dearden, J. C. Eur. J. Med. Chem. 2009, 44, 473.
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