Novel 4-thiazolidinone derivatives as potential antifungal and antibacterial drugs

Article abstract of DOI:10.1016/j.bmc.2009.10.041

As part of ongoing studies in developing new antimicrobials, a class of structurally novel 4-thiazolidinone derivatives incorporating three known bioactive nuclei such as thiazole, thiazolidinone and adamantane was synthesized by the multi-step reaction protocol, already reported in the literature. NMR and Molecular Modeling techniques were employed for structure elucidation and Z/E potential isomerism configuration of the analogues. Evaluation of antibacterial and antifungal activity showed that almost all compounds exhibited better results than reference drugs thus they could be promising candidates for novel drugs.

Full text of DOI:10.1016/j.bmc.2009.10.041

Bioorganic & Medicinal Chemistry 18 (2010) 426–432
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Novel 4-thiazolidinone derivatives as potential antifungal
and antibacterial drugs
b
c
Kouatli Omar a, Athina Geronikaki a, , Panagiotis Zoumpoulakis , Charalabos Camoutsis ,
*
d
d
d
´
´
´
ˇ
Marina Sokovic , Ana Ciric , Jasmina Glamoclija
a School of Pharmacy, Department of Pharmaceutical Chemistry of Aristotle, University of Thessaloniki, Thessaloniki, Greece
b Laboratory of Molecular Analysis, Institute of Organic and Pharmaceutical Chemistry, National Hellenic Research Foundation, 48 Vas. Constantinou Avenue, 11635 Athens, Greece
c School of Health Sciences, Department of Pharmacy, Laboratory of Pharmaceutical Chemistry, University of Patras, Patras, Greece
d Department of Plant Physiology, Mycological Laboratory, Institute of Biological Research, Bulevar Despota Stefana 142, 11000 Belgrade, Serbia
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 1 July 2009
Accepted 23 October 2009
Available online 27 October 2009
As part of ongoing studies in developing new antimicrobials, a class of structurally novel 4-thiazolidinone
derivatives incorporating three known bioactive nuclei such as thiazole, thiazolidinone and adamantane
was synthesized by the multi-step reaction protocol, already reported in the literature. NMR and Molec-
ular Modeling techniques were employed for structure elucidation and Z/E potential isomerism configu-
ration of the analogues. Evaluation of antibacterial and antifungal activity showed that almost all
compounds exhibited better results than reference drugs thus they could be promising candidates for
novel drugs.
Keywords:
4-Thiazolidinone
Antibacterial
Antifungal
Ó 2009 Elsevier Ltd. All rights reserved.
Z/E isomerism
1. Introduction
2. Results and discussion
The treatment of infectious diseases still remains an important
and challenging problem because of a combination of factors
including emerging infectious diseases and the increasing number
of multi-drug resistant microbial pathogens with particular rele-
vance for Gram positive bacteria.1–5 On the other hand, a recent
survey of novel small-molecule therapeutics revealed that the
majority of them result from an analogue-based approach and that
their market value represents two-thirds of all drug sales.6
Because of this, and given our recent finding of a new class of anti-
bacterial agents, the 2-thiazolylimino-5-benzylidene-4-thiazolidi-
nones7,8 we decided to extend our research to classes of analogues.
It was found that 4-thiazolidinone ring system which is a core struc-
ture in various synthetic pharmaceuticals display a broad spectrum
of biological activity, including antibacterial and antifungal proper-
ties.9–14 Adamantane derivative have long been known for their
antiviral activity against Influenza A15–20 and HIV viruses.21,22 Sev-
eral adamantane derivatives were also associated with central ner-
vous,23–25 antimicrobial,26–30 and anti-inflammatory activities.31
Our analogue-based design encompasses the synthesis of ele-
ven new of 4-adamantyl-2-thiazolylimino-5-arylidene-4-thiazo-
lidinone 5a–k derivatives (Fig. 1), to be tested for their in vitro
antimicrobial properties against Gram positive and Gram negative
bacteria and fungi.
2.1. Chemistry
As part of our ongoing studies in developing new antimicrobi-
als7,8 we report the synthesis of new class of structurally novel
4-thiazolidinone derivatives incorporating three known bioactive
nuclei such as thiazole, thiazolidinone and adamantane.
R
S
N
O
N
N
S
H
5a: R = 4-Cl
5b: R = 3-Cl
5c: R = 2-Cl
5d: R = 4-NO2
5e: R = 3-NO2
5f: R = 2-NO2
5g: R = 4-OH
5h: R = 4-OH, 3-OCH3
5i: R = 4-OH, 3,5-OCH3
5j: R = 4-OCH3
* 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 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2009.10.041

K. Omar et al. / Bioorg. Med. Chem. 18 (2010) 426–432
427
The compounds described in this paper were synthesized by the
Compound 4 inhibited fungal growth at 0.60–2.38 Â 10À2
mlwhilefungicidalactivitywas achieved at 2.38–3.35 Â 10À2
l
l
mol/
mol/
multi-step reaction protocol reported earlier by us7 (Scheme 1). 2-
Chloracetylchloride of 4-adamantyl-2-aminothiazole (3), synthe-
sized using procedure reported earlier32 from 4-adamantyl-2-ami-
nothiazole (2), was undergone heterocyclization in the presence of
ammonium isothiocyanate in refluxing ethanol33,7 producing 2-(4-
adamantylthiazol-2-ylimino)thiazolidin-4-one (4). The 4-adaman-
tyl-2-thiazolylimino-5-arylidene-4-thiazolidinones 5a–j were ob-
tained by refluxing 4 with appropriate aldehydes in buffered
glacial acetic acid (Scheme 1).
ml. This compound showed the lowest antifungal activity, expressed
as minimal inhibitory concentration (MIC) against Penicillium funi-
culosum (MIC 1.79 Â 10À2
lmol/ml) Aspergillus flavus (MIC
1.79 Â 10À2
mol/ml) and Aspergillus versicolor (MIC 2.38 Â
l
10À2
lmol/ml), moderate activity against Trichoderma viride, Asper-
gillus fumigatus, Aspergillus niger with MIC 1.19 Â 10À2
lmol/ml,
whereas it exhibited a strong effectiveness towards Penicillium
ochrochloron and Fulvia fulvum (MIC 0.60 Â 10À2
lmol/ml). In all
All the new compounds 5a–j were characterized by mp, elemen-
tal analyses and spectroscopic data (1H NMR, MS and IR). The spec-
tral data and the elemental analysis of the new compounds reported
in this study correlate with the proposed structures.
cases activity of compound 4 was better than activity of two refer-
ence drugs, bifonazole and ketoconazole.
More significant inhibitory properties were detected for 5-ary-
lidene derivatives 5a–j.
The mechanism of the cyclocondensation step, the amino–imi-
no tautomeric equilibrium of the heteroarylthiazolidinones and of
their adamantane derivatives 5a–5j, the E/Z potential isomerism of
the latter, were the same that was investigated through the analy-
sis of IR and 1H NMR spectral data for our compounds reported
previously7,8 and confirmed on the basis of the literature data.34
All the compounds tested showed fungistatic activity at 0.52–
2.38 Â 10À2
l
mol/ml and fungicidal at 1.05–3.35 Â 10À2
lmol/ml
against all the fungi tested. Derivatives 5a–5c exhibited fungistatic
effect at 0.55–2.19 Â 10À2
l
mol/ml and fungicidal activity was ob-
mol/ml. In this group compound 5b
served at 1.64–3.23 Â 10À2
l
showed the best antifungal potential. Compounds 5d–5f possessed
almost the same activity, MIC at 0.54–2.15 Â 10À2
lmol/ml, and
2.2. Antibacterial/antifungal activity
MFC 1.08–2.15 Â 10À2
lmol/ml. Derivatives 5g–5j showed MIC at
0.52–1.73 Â 10À2
l
mol/ml and MFC at 1.05–2.31 Â 10À2
lmol/ml,
The results of antifungal activity of 4-[(adamantan-1-yl)1,3-
(thiazol-2-ylimino)] thiazolidin-4-one 4 and its 5-arylidene deriv-
atives compounds 5a–5j against a panel of selected Gram positive,
Gram negative bacteria and fungi are presented in Table 1 in com-
parison with those of the reference drugs ampicillin and strepto-
mycin, bifonazole and ketoconazole, respectively.
where compound 5i exhibited the highest antifungal potential
with MIC at 0.52–1.57 Â 10À2
lmol/ml and MFC at 1.05–2.09. This
compound showed the best antifungal effect among all the tested.
The majority of compounds showed the worst activity against
A. versicolor, while F. fulvum is the most sensitive species. The most
active compounds against this species (F. fulvum) among all tested
are compounds 5i (0.52 Â 10À2
lmol/ml), followed by 5f and 4.
Taking into account that almost all compounds exhibited activ-
ity better than reference drugs, they could be promising candidates
for antifungal drugs.
In addition compound 5a–j were evaluated for antibacterial
activity against a wide number of Gram positive bacteria, as well
as Gram negative bacteria. The antibacterial activity of compounds
tested by microdilution method, are presented in Table 2. The kind
of the exerted antibacterial activity was investigatedby determining
the minimal bactericidal concentrations (MBCs). The experimental
data (second values) presented in Table 2 show that 4-adamantyl-
2-thiazolylimino-5-arylidene-4-thiazolidinones 5a–j possess bac-
teriostatic properties, being MBCs almost twofold higher than the
corresponding MICs.
N
a
+
H2NCSNH2
COCH2Br
NH2
1
S
2
O
b
d
c
N
N
HN
S
NHCOCH2Cl
N
S
S
All compounds showed strong antibacterial activity against all
bacterial species. MIC for compounds 5a–5j is at 1.05–
3
4
R
4.76 Â 10À2
l
mol/ml and MBC at 1.57–7.17 Â 10À2
lmol/ml. Com-
pound 4 (initial thiazolidinone) exhibited the lowest antibacterial
activity among all the other tested, with MIC 1.79–
2.38 Â 10À2
l
mol/ml and MBC at 2.38–7.17 Â 10À2
lmol/ml.
S
N
Group of compounds 5a–5c showed MIC at 1.10–
4.38 Â 10À2
l
mol/ml and MBC 2.19–6.57 Â 10À2
lmol/ml, where
O
N
N
S
5b had the best activity. MIC of compounds 5d–5f is at 1.08–
H
4.30 Â 10À2
l
mol/ml and MBC at 2.15–6.45 Â 10À2
lmol/ml. These
5
compounds showed almost the same activity. Among derivatives
5a: R = 4-Cl
5b: R = 3-Cl
5c: R = 2-Cl
5d: R = 4-NO2
5e: R = 3-NO2
5f: R = 2-NO2
5g: R = 4-OH
5g–5j (MIC at 1.05–4.62 Â 10À2
lmol/ml and MBC 1.57–
6.93 Â 10À2
lmol/ml) compound 5j possessed the best antibacte-
rial activity, MIC 1.05–4.18 Â 10À2
lmol/ml and MBC at 2.09–
6.27 Â 10À2
lmol/ml. It can be seen that this compound in general
showed the highest antibacterial activity. More significant inhibi-
tory properties were detected for compound 5i against Micrococcus
flavus (ATCC 10240), Staphylococcus aureus as well as towards Sal-
5h: R = 4-OH, 3-OCH3
5i: R = 4-OH, 3,5-OCH3
5j: R = 4-OCH3
monella typhimurium (MICs 1.05 Â 10À2
lmol/ml).
The most sensitive bacterial species on compounds tested is
Gram positive bacteria, especially, Bacillus cereus, while Gram neg-
ative bacteria Pseudomonas mirabilis is the most resistant species. It
can be seen that all the compounds tested on antibacterial activity
Scheme 1. Synthesis of the compounds. Reagents and conditions: (a) isopropanol,
stirring 0.5 h, rt: (b) ClCOCH2Cl, N,NDMF, rt, 2 h; (c) NH4SCN, EtOH, reflux, 1–2 h; (d)
RC6H4CHO, MeCOOH, MeCOONa.

428
K. Omar et al. / Bioorg. Med. Chem. 18 (2010) 426–432
Table 1
Antifungal activity of tested compounds and fungicides (MIC and MFC in
l
mol/ml  10À2
)
Fungi
4
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
Bif
Ket
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
MIC
MFC
P. funiculosum
P. ochrochloron
T. viride
1.79
2.38
1.10
2.19
2.19
2.19
2.19
3.23
2.15
2.15
1.08
2.15
1.08
2.15
1.72
2.29
1.64
2.19
1.05
2.09
1.16
2.31
64.0
80.0
38.0
95.0
0.60
2.38
1.64
2.19
0.55
1.10
1.10
2.19
1.61
2.15
1.08
2.15
1.08
2.15
1.72
2.29
1.64
2.19
0.52
1.05
1.16
2.31
48.0
64.0
380.0
380.0
1.19
2.38
1.10
1.64
1.10
2.19
1.10
2.19
1.08
2.15
0.54
1.08
1.08
2.15
1.15
2.29
1.10
2.19
1.05
2.09
1.16
2.31
64.0
80.0
475.0
570.0
A. fumigatus
A. niger
1.19
2.38
1.10
2.19
1.10
2.19
1.64
2.19
1.61
2.15
1.61
2.15
1.08
2.15
1.15
2.29
1.64
2.19
1.57
2.09
1.73
2.31
48.0
64.0
38.0
95.0
1.19
2.38
1.10
2.19
1.10
2.19
1.10
2.19
1.08
2.15
1.08
2.15
1.61
2.15
1.15
2.29
1.64
2.19
1.05
2.09
1.73
2.31
48.0
64.0
38.0
95.0
A. flavus
1.79
2.38
1.10
2.19
1.64
2.19
1.10
2.19
1.61
2.15
1.61
2.15
1.61
2.15
1.15
2.29
1.64
2.19
1.05
2.09
1.73
2.31
48.0
64.0
285.0
380.0
A. versicolor
F. fulvum
2.38
3.35
1.64
2.19
1.10
2.19
1.64
2.19
1.61
2.15
1.61
2.15
1.08
1.61
1.72
2.29
1.64
2.19
1.57
2.09
1.73
2.31
32.0
64.0
38.0
95.0
0.60
1.19
1.10
1.64
1.64
2.19
1.10
1.64
1.61
2.15
1.08
1.61
0.54
1.08
1.15
2.29
1.10
2.19
0.52
2.09
0.58
2.31
32.0
64.0
38.0
95.0
Table 2
Antibacterial activity of tested compounds and antibiotics (MIC and MBC in
l
mol/ml  10À2
)
Bacteria
4
MIC
MBC
5a
MIC
MBC
5b
MIC
MBC
5c
MIC
MBC
5d
MIC
MBC
5e
MIC
MBC
5f
MIC
MBC
5g
MIC
MBC
5h
MIC
MBC
5i
MIC
MBC
5j
MIC
MBC
Str
MIC
MBC
Amp
MIC
MBC
B. cereus
1.79
2.38
1.64
2.19
1.64
2.19
2.19
4.38
2.15
2.15
1.61
2.15
1.08
2.15
1.72
2.29
1.10
2.19
2.09
2.09
2.31
2.31
4.3
8.6
24.8
37.2
M. flavus
2.38
4.76
1.10
2.19
1.10
2.19
1.64
2.38
2.15
2.15
2.15
4.30
2.15
4.30
2.29
4.58
1.10
2.19
1.05
2.09
2.31
4.62
8.6
17.2
24.8
37.2
S. aureus
2.38
4.76
2.19
4.38
1.64
4.38
2.19
4.38
1.61
4.30
2.15
4.30
1.61
2.15
1.72
4.58
2.19
4.38
1.05
1.57
2.31
4.62
17.2
34.4
24.8
37.2
E. coli
2.38
4.76
2.19
4.38
2.19
4.38
2.19
4.38
2.15
4.30
2.15
2.15
2.15
4.30
2.29
4.58
1.10
1.64
2.09
4.18
1.16
2.31
17.2
34.4
37.2
49.2
P. aeruginosa
P. mirabilis
S. typhimuirium
L. monocyto
1.79
4.76
2.19
4.38
2.19
4.38
2.19
4.38
2.15
4.30
2.15
4.30
2.15
4.30
2.29
4.58
1.64
4.38
1.57
4.18
1.16
2.31
17.2
34.4
74.4
124.0
4.76
7.17
4.38
6.57
4.38
6.57
4.38
6.37
4.30
6.45
4.30
6.45
4.30
6.45
4.58
6.87
4.38
6.57
4.18
6.27
4.62
6.93
17.2
34.4
37.2
49.2
2.38
4.76
2.19
4.38
2.19
4.38
2.19
4.38
2.15
2.15
1.08
2.15
2.15
4.30
1.15
1.72
2.19
4.38
1.05
1.57
1.16
2.31
17.2
34.4
24.8
49.2
2.38
4.76
2.19
4.38
2.19
4.38
2.19
4.38
2.15
4.30
2.15
4.30
2.15
4.30
2.29
4.58
1.10
2.19
2.09
4.18
2.31
4.62
25.8
51.6
37.2
74.4
showed much better effect than commercial antibiotics, strepto-
carrying hydrophilic hydroxyl or methoxy group (5g, 5j) or nitro
substituted compounds (5d–f).
mycin and ampicillin (MIC 4.3–25.8 Â 10À2
lmol/m and MBC
8.6–51.6 Â 10À2
l
mol/ml—for streptomycin, and MIC 24.8–
It is interesting to point out that for the isomeric chloro
substituted compounds (5a–c) the meta substituted derivative
(5b) is mostly endowed with higher activity with respect to
ortho and para (5a and 5c) while for nitro substituted com-
pounds 5d–f the most active are para and meta (5d and 5e)
derivatives.
The introduction of methoxy group in position 3 and 3, 5 (5h–i)
in the 4-hydroxy derivative (5g) in general lead to compounds with
higher activity, whereas replacement of hydroxyl group with
methoxy in the 4-position in most cases has the opposite effect.
Only in case of Gram negative bacteria P. aeruginosa the replace-
ment of hydroxy group in position 4 with methoxy (5j) enlarge
the antimicrobial activity.
74.4 Â 10À2
l
mol/ml and MBC 37.2–124.0 Â 10À2
lmol/ml—for
ampicillin). Thus all these compounds could be used as lead com-
pounds for new antibacterial drugs.
Compounds 5i in general showed the highest antibacterial as
well as antifungal activity, while compound 4 exhibited the lowest
antimicrobial potential.
It has been known that the introduction of arylidene moieties at
different positions of the thiazolidinone ring enhanced the antimi-
crobial activity.7,35,36 As regards the relationships between the
structure and the detected antibacterial activity, the 5-arylidene
derivatives 5a–j showed a significant antibacterial efficacy, greater
than that of parent 4-[adamantyl-2-(thiazol-2-ylimino)] thiazoli-
din-4-one 4 and appear to be dependent on the substitution at
the benzene ring as well.
Almost the same behavior was observed for antifungal activity
of tested compounds.
Thus, the chloro derivatives (5a–c) were found to be more effec-
tive against all tested microorganisms than arylidene derivatives
As a conclusion it can be noticed that fungi were in general
more sensitive than bacterial species on these compounds.

K. Omar et al. / Bioorg. Med. Chem. 18 (2010) 426–432
429
4 h and then poured into ice-cold water. The precipitate was fil-
tered and washed with water and the resulting crude product
was purified by recrystallisation from dioxane.
3. Conclusion
The newly synthesized 4-adamantyl-2-thiazolylimino-5-arylid-
ene-4-thiazolidinones 5a–j, exhibit a remarkable inhibition of the
growth of a wide spectrum of Gram positive bacteria, Gram nega-
tive bacteria and fungi. The most sensitive bacterial species on
compounds tested is Gram positive bacteria, B. cereus, while Gram
negative bacteria Proteus mirabilis is the most resistant species. As far
as concern the fungi, the majority of compounds showed the worst
activity against A. versicolor, while F. flavum is the most sensitive
species.
It should be noticed that all compounds tested exhibited better
activity than commercial antimicrobial agents used as reference
drugs and few times higher activity than ketoconazole.
All the compounds 5a–j showed excellent antibacterial activ-
ity indicating that the diverse substitutions were well tolerated
on the benzylidene moiety for proper fit at the potential receptor
site.
4.1.4.1. (2Z,5Z)-2-{[4-(Adamantan-1-yl)-1,3-thiazol-2-yl]i-
mino}-5-(4-chlorobenzylidene)-1,3-thiazolidin-4-one
(5a). Yield: 76%. Mp 270–270.5 °C (dioxane). TLC: eluent = ben-
zene–ethanol (9:1), Rf = 0.73. IR
m
cmÀ1: 3075 (NH), 1720 (C@O),
1592 (C@N). 1H NMR (DMSO-d6) d (ppm): 1.71–2.07 (m, 15H,
adam.), 6.95 (s, 1H, thiazolyl), 7.55 (d, 2H, ArH, J = 9 Hz), 7.66 (m,
2H, ArH and 1H, @CH), 12.63 (s, 1H, NH). Anal. Calcd for
C23H22N3OS2Cl (MW 455.5): C, 60.59; H, 4.82; N, 9.22. Found: C,
60.54; H, 4.80; N, 9.24.
4.1.4.2. (2Z,5Z)-2-{[4-(Adamantan-1-yl)-1,3-thiazol-2-yl]i-
mino}-5-(3-chlorobenzylidene)-1,3-thiazolidin-4-one
(5b). Yield: 86%. Mp 263–265 °C (dioxane). TLC: eluent = benzene–
ethanol (9:1), Rf = 0.76. IR
m
cmÀ1: 3090 (NH), 1710 (C@O), 1592
(C@N). 1H NMR (DMSO-d6) d (ppm): 1.75–2.05 (m, 15H, adam.),
6.96 (s, 1H, thiazolyl), 7.51 (d, 2H, ArH, J = 8.3 Hz), 7.65 (m, 2H,
ArH and 1H, @CH), 12.68 (s, 1H, NH). Anal. Calcd for C23H22N3OS2Cl
(MW 455.5): C, 60.59; H, 4.82; N, 9.22. Found: C, 60.55; H, 4.83; N,
9.23.
The outstanding properties of this new class of antibacterial sub-
stances deserve further investigation in order to clarify the mode of
action at molecular level, responsible for the activity observed.
4. Experimental
4.1.4.3. (2Z,5Z)-2-{[4-(Adamantan-1-yl)- 1,3-thiazol-2-yl]imino}-
5-(2-chlorobenzylidene)-1,3-thiazolidin-4-one (5c). Yield: 70%.
Mp 214–215 °C (dioxane). TLC: eluent = benzene–ethanol (9:1),
4.1. Chemistry—general aspects
Melting points were taken in glass capillary tubes on a Haake
Bucher apparatus and are uncorrected. IR spectra were recorded
on a FT-IR Jasco spectrophotometer in solid phase KBr. All proton
NMR spectra were determined with a Varian 300 MHz spectrome-
ter using deuterated dimethylsulfoxide (DMSO-d6) and are re-
ported in d (ppm) units. Thin layer chromatography (TLC) was
performed in E. Merck precoated silica gel plates. Visualization
was obtained by exposure to iodine vapors and/or under UV light
(254 nm). The elemental analyses (C, H, N) of all compounds were
performed by the Center of Instrumental Analysis of the University
of Patras.
Rf = 0.64. IR
m
cmÀ1: 3100 (NH), 1720 (C@O), 1598 (C@N). 1H NMR
(DMSO-d6) d (ppm): 1.64–2.00 (m, 15H, adam.), 6.93 (s, 1H, thiazol-
yl), 7.42 (m, 1H, ArH), 7.49 (m, 1H, ArH), 7.63 (d, 1H, ArH, J = 8 Hz),
7.71 (d, 1H, ArH, J = 7.8 Hz), 7. 97 (s, 1H, @CH), 12.71 (s, 1H, NH).
Anal. Calcd for C23H22N3OS2Cl (MW 455.5): C, 60.59; H, 4.82; N,
9.22. Found: C, 60.65; H, 4.81; N, 9.21.
4.1.4.4. (2Z,5Z)-2-{[4-(Adamantan-1-yl)-1,3-thiazol-2-yl]i-
mino}-5-(4-nitrobenzylidene)-1,3-thiazolidin-4-one (5d). Yield:
64%. Mp 285–287 °C (dioxane). TLC: eluent = benzene–ethanol
(9:1), Rf = 0.74. IR
m
cmÀ1: 3080 (NH), 1715 (C@O), 1599 (C@N).
1H NMR (DMSO-d6) d (ppm): 1.70–2.07 (m, 15H, adam.), 6.95 (s,
1H, thiazolyl), 7.75 (s, 1H, @CH), 7.88 (d, 2H, ArH, J = 8.6 Hz), 8.24
(d, 2H, ArH, J = 8.7 Hz), 12.77 (s, 1H, NH). MS: 465 (M+, 100%),
331 (72), 310 (4), 280 (9), 264 (5), 257 (55), 240 (20), 226 (3), 98
(7), 88 (9), 78 (11), 74 (4), 61 (6). Anal. Calcd for C23H22N4O3S2
(MW 466): C, 59.22; H, 4.72; N, 12.01. Found: C, 59.29; H, 4.71;
N, 12.03.
4.1.1. Synthesis of 4-adamantyl-2-aminothiazole (1)
To a solution of 1-adamantyl bromomethyl ketone, 1, (257 mg,
1 mmol) in 5 ml of isopropanol, a suspension of thiourea (152 mg,
2 mmol) in 10 ml of isopropanol was added. The mixture was stir-
red for half an hour. After this time the resulting solution was
poured into a solution of sodium carbonate, and the precipitate
formed was filtered and dried to give, after recrystallization from
ethylacetate, 220 mg (94%) of pure product, 2. Mp 215–215.5 °C.
4.1.4.5. (2Z,5Z)-2-{[4-(Adamantan-1-yl)-1,3-thiazol-2-yl]i-
mino}-5-(3-nitrobenzylidene)-1,3-thiazolidin-4-one (5e). Yield:
54%. Mp 264–265 °C (dioxane). TLC: eluent = benzene–ethanol
IR (KBr):
m
= 3050 cmÀ1 (N–H).
4.1.2. Synthesis of chloroacetylchloride of 4-adamantyl-2-
aminothiazole
(9:1), Rf = 0.85. IR
m
cmÀ1: 3085 (NH), 1700 (C@O), 1599 (C@N).
1H NMR (DMSO-d6) d (ppm): 1.73–2.04 (m, 15H, adam.), 6.97 (s,
1H, thiazolyl), 7.78 (t, 1H, ArH, J = 8 Hz), 7.81 (s, 1H, @CH), 8.10
(d, 1H, ArH, J = 8.5 Hz), 8.27 (d, 1H, ArH, J = 8.5 Hz), 8.47 (s, 1H,
ArH), 12.74 (s, 1H, NH). Anal. Calcd for C23H22N4O3S2 (MW 466):
C, 59.22; H, 4.72; N, 12.01. Found: C, 59.15; H, 4.71; N, 12.00.
It was performed according to previous described method.7
4.1.3. Synthesis of 4-adamantyl-2-(thiazol-2-ylimino)
thiazolidin-4-one (4)
A solution of 2-chloro-N-(4-adamnatyl thiazole) acetamide
(5 mmol) and ammonium thiocyanate (10 mmol) in 20 mL of 96%
ethanol was refluxed for 1 h and allowed to stand overnight. The
precipitate was filtered washed with water and then recrystallised.
4.1.4.6. 2-{[4-(Adamantan-1-yl)-1,3-thiazol-2-yl]imino}-5-(2-
nitrobenzylidene)-1,3-thiazolidin-4-one (5f). Yield: 68%. Mp
208–210 °C (dioxane). TLC: eluent = benzene–ethanol (9:1),
Rf = 0.80. IR
m
cmÀ1: 3087 (NH), 1718 (C@O), 1600 (C@N). 1H
4.1.4. General procedure for synthesis of 4-adamantyl-2-
thiazolylimino-5-arylidene-4-thiazolidinones 5a–j
A well-stirred solution of 0.8 g of 2-(4-adamantylthiazol-2-yli-
mino) thiazolidin-4-one (4 mmol) in 35 ML of glacial acetic acid
was buffered with sodium acetate (8 mmol) and added with the
appropriate arylaldehyde (6 mmol). The solution was refluxed for
NMR (DMSO-d6) d (ppm): 1.70–2.00 (m, 15H, adam.), 6.82, 6.92
(ss, 1H, thiazolyl), 7.88 (s, 1H, @CH), 7.71 (m, 1H, ArH), 7.82
(m, 1H, ArH), 7.84 (d, 1H, ArH, J = 7.6 Hz), 8.18 (d, 1H, ArH,
J = 8 Hz), 11.95,12.71 (ss, 1H, NH). Anal. Calcd for C23H22N4O3S2
(MW 466): C, 59.22; H, 4.72; N, 12.01. Found: C, 59.25; H, 4.73;
N, 12.02.

430
K. Omar et al. / Bioorg. Med. Chem. 18 (2010) 426–432
4.1.4.7. (2Z,5Z)-2-{[4-(Adamantan-1-yl)-1,3-thiazol-2-yl]i-
mino}-5-(4-hydroxybenzylidene)-1,3-thiazolidin-4-one
(5g). Yield: 38%. Mp 250–251 °C (dioxane). TLC: eluent = benzene–
microplates were incubated for 72 h at 28 °C, respectively. The
lowest concentrations without visible growth (at the binocular
microscope) were defined as MICs.
The fungicidal concentrations (MFCs) were determined by serial
sub-cultivation of a 2 ll into microtiter plates containing 100 ll of
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, Belgarde,
Serbia) and ketoconazole (Zorkapharma, Šabac, Serbia), were used
ethanol (9:1), Rf = 0.72. IR
m
cmÀ1: 3500 (OH), 3085 (NH), 1700
(C@O), 1580 (C@N). 1H NMR (DMSO-d6) d (ppm): 1.62–1.98 (m,
15H, adam.), 6.78–6.97 (m, 3H, ArH and thiazolyl), 7.46–7.50 (m,
3H, ArH and @CH), 10.24 (s, 1H, OH), 12.41 (s, 1H, NH). Anal. Calcd
for C23H23N3O2S2 (MW 437): C, 63.15; H, 5.26; N, 9.61. Found: C,
63.20; H, 5.28; N, 9.62.
4.1.4.8. (2Z,5Z)-2-{[4-(Adamantan-1-yl)-1,3-thiazol-2-yl]i-
mino}-5-(4-hydroxy-3-methoxy-benzylidene)-1,3-thiazolidin-
4-one (5h). Yield: 32%. Mp 251–252 °C (dioxane). TLC: elu-
as positive controls (1–3000 lg/ml). All experiments were per-
formed in duplicate and repeated three times.
ent = benzene–ethanol (9:1), Rf = 0.72. IR
m
cmÀ1: 3550 (OH),
4.2.2. Test for antibacterial activity
3085 (NH), 1699 (C@O), 1589 (C@N). 1H NMR (DMSO-d6) d
(ppm): 1.70–2.00 (m, 15H, adam.), 3.81 (s, 3H, OCH3), 6.86 (d,
1H, ArH, J = 8.2 Hz), 6.91 (s, 1H, thiazolyl), 7.15 (d, 1H, ArH,
J = 8.3 Hz), 7.20 (s, 1H, ArH), 7.59 (s, 1H, @CH), 9.92 (s, 1H,
OH),12.46 (s, 1H, NH). Anal. Calcd for C24H25N3O3S2 (MW 467): C,
61.67; H, 5.35; N, 8.99. Found: C, 61.62; H, 5.37; N, 9.00.
The following Gram negative bacteria were used: Escherichia
coli (ATCC 35210), Pseudomonas aeruginosa (ATCC 27853), S.
typhimurium (ATCC 13311), Proteus mirabilis (human isolate) and
the following Gram positive bacteria: B. cereus (clinical isolate),
M. flavus (ATCC 10240), Listeria monocytogenes (NCTC 7973), and
S. aureus (ATCC 6538). The organisms were obtained from the
Mycological Laboratory, Department of Plant Physiology, Institute
´
4.1.4.9. (2Z,5Z)-2-{[4-(Adamantan-1-yl)-1,3-thiazol-2-yl]i-
mino}-5-(4-hydroxy-3,5-dimethoxy-benzylidene)-1,3-thiazoli-
din-4- (5i). Yield: 49%. Mp 263–263.5 °C (dioxane). TLC:
for Biological Research ‘Siniša Stankovic’, Belgrade, Serbia.
The antibacterial assay was carried out by microdilution meth-
od37–39 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 Â 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. All experiments were per-
formed in duplicate and repeated three times.
eluent = benzene–ethanol (9:1), Rf = 0.67. IR
m
cmÀ1: 3450 (OH),
3085 (NH), 1700 (C@O), 1594 (C@N). 1H NMR (DMSO-d6) d
(ppm): 1.69–2.00 (m, 15H, adam.), 3.81 (s, 6H, OCH3), 6.93 (s, 1H,
thiazolyl), 6.95 (s, 2H, ArH), 7.62 (s, 1H, @CH), 9.29 (s, 1H, OH),
12.47 (s, 1H, NH). Anal. Calcd for C25H27N3O4S2 (MW 497): C,
60.36; H, 5.43; N, 8.45. Found: C, 60.41; H, 5.41; N, 8.47.
4.1.4.10. (2Z,5Z)-2-{[4-(Adamantan-1-yl)-1,3-thiazol-2-yl]i-
mino}-5-(4-methoxy-benzylidene)-1,3-thiazolidin-4-one
(5j). Yield: 43%. Mp 260–260.5 °C (dioxane). TLC: eluent = ben-
4.2.3. Microdilution test
The minimum inhibitory and bactericidal concentrations (MICs
and MBCs) were determined using 96-well microtitre plates. The
bacterial suspension was adjusted with sterile saline to a concen-
tration of 1.0 Â 105 cfu/ml. Compounds to be investigated were
zene–ethanol (9:1), Rf = 0.61. IR
m
cmÀ1: 3100 (NH), 1710 (C@O),
1598 (C@N). 1H NMR (DMSO-d6) d (ppm): 1.71–2.05 (m, 15H,
adam.), 3.81 (s, 3H, OCH3), 6.91 (s, 1H, thiazolyl), 7.01 (d, 2H,
ArH, J = 8.7 Hz), 7.64 (m, 2H, ArH and 1H, @CH), 12.5 (s, 1H, NH).
Anal. Calcd for C24H25N3O2S2 (MW 451): C, 63.85; H, 5.54; N,
9.31. Found: C, 63.91; H, 5.56; N, 9.34.
dissolved in broth LB medium (100 ll) with bacterial inoculum
(1.0 Â 104 cfu per well) to achieve the wanted concentrations
(1 mg/ml). The microplates were incubated for 24 h at 48 °C. The
lowest concentrations without visible growth (at the binocular
microscope) were defined as concentrations that completely inhib-
ited bacterial growth (MICs). The MBCs were determined by serial
4.2. Biological evaluation
4.2.1. Test for antifungal activity
sub-cultivation of 2 ll into microtitre plates containing 100 ll of
For the antifungal bioassays, eight fungi were used: A. flavus
(ATCC 9643), A. fumigatus (plant isolate), A. niger (ATCC 6275), A.
versicolor (ATCC 11730), F. fulvum (TK 5318), P. funiculosum (ATCC
36839), P. ochrochloron (ATCC 9112) and T. viride (IAM 5061). The
organisms were obtained from the Mycological Laboratory,
Department of Plant Physiology, Institute for Biological Research
broth per well and further incubation for 72 h. The lowest concen-
tration 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 man-
ager 4.0 (Bio-Rad Laboratories) and compared with a blank and the
positive control. Streptomycin (Sigma P 7794) and Ampicillin
(Panfarma, Belgarde, Serbia) were used as a positive control
(1 mg/ml DMSO). All experiments were performed in duplicate
and repeated three times.
´
‘Siniša Stankovic’, Belgrade, Serbia.
The micromycetes were maintained on malt agar and the
cultures stored at 4 °C and sub-cultured once a month.37 In order
toinvestigatetheantifungalactivityoftheextracts, amodifiedmicro
dilution technique was used.38–40 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 Â 105 in a final
4.3. NMR spectroscopy
Ultra precision NMR tubes Wilmad 535–5 mm were used for
the NMR experiments. Compounds 5a–5j were dissolved in
DMSO-d6 and a series of experiments were performed using Varian
600 MHz spectrometer at 300 K. All data are collected using pulse
sequences and phase-cycling routines provided in the Varian
libraries. The 1H spectral width was set to 8500 Hz at 600 MHz.
Typically the 2D NOESY spectrum was acquired with 4096 data
points in t2 dimension, 64 scans, 256 increments in t1 dimension,
150 ms mixing time and a relaxation delay of 1 s. Data processing
including apodization with cosine square bell function, Fourier
volume of 100 ll per well. The inocula were stored at 4 °C for further
use. Dilutions of the inocula were cultured on solid malt agar to
verify the absence of contamination and to check the validity 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 DMSO
(1 mg/ml) and added in broth Malt medium with inoculum. The

K. Omar et al. / Bioorg. Med. Chem. 18 (2010) 426–432
431
transformation and phasing, were performed using Varian VNMR
software.
4.4. Molecular modeling
Computer calculations were performed using Quanta software
of Molecular Simulations on a Silicon Graphics O2. Molecular
Mechanics calculations were carried out using the CHARMm force
field. Initial 3D structures were first minimized with steepest des-
cents and then with Newton-Raphson algorithms using an energy
tolerance of 0.01 kcal/molÀ1 AÀ1, to reach a local minimum. The
dielectric constant (e) was set to 45 during minimization simulat-
ing the DMSO environment. To generate conformers, a random
search procedure (random sampling) was applied. This method
randomly changes all defined torsion angles within a predefined
angular window. The range of the torsion angle varies during the
search procedure and generates new conformations. CHARMm en-
ergy minimization is performed for each randomly altered confor-
mation. In order to explore the preferred torsion angles that
correspond to the lowest energy conformers and energy barriers
of the molecules under study, stochastic search procedures (sys-
tematic grid scan) were used. This method initiates a grid scan
search that generates conformations by varying specified torsion
angles over a grid of equally spaced values. Intervals of 5° were ap-
plied for the rotation of one bond and 10° for simultaneous rota-
tion of two bonds. During these searches, the predetermined
torsion angle remained constant while minimization using 1000
steps of conjugate gradient algorithm was applied to ‘relax’ the
whole molecule.
Figure 3. Representative low energy conformers of 2-{[4-(adamantan-1-yl)- 1,3-
thiazol-2-yl]imino}-5-(4-nitrobenzylidene)-1,3-thiazolidin-4-one derived from
Random Sampling conformational search, after clustering by torsions. Only 2Z,5Z
configuration can satisfy the observed NOEs between adamantine and para-
substituted aromatic ring protons.
proton resonated, as expected, at higher chemical shift values
due to the deshielding effect of the adjacent C@O, than it would
do in E configuration isomers, because of the lower deshielding ef-
fect of 1-S as already was mentioned in our previous paper,7,8 as
well as in the literature.34,35
4.5. Stereochemistry and conformational properties
Regarding the stereochemistry of the C@N imino exocyclic dou-
ble bond, 2D NOESY NMR spectroscopy has shown NOE signals be-
tween adamantane and aromatic ring protons (Fig. 2). This
experimental result suggests that these two moieties are close in
space. Based on this finding, we proceeded with Molecular Model-
ing and conformational analysis studies in order to find geometries
of the molecules to satisfy the observed NOEs. In conclusion, only
2Z,5Z isomers can satisfy this distance constrain which brings in
proximity the adamantane and the aromatic ring (Fig. 3).
Assignment of the NH peak at ꢀ12.5 ppm for the Z isomers on
the C@N imino exocyclic double bond, results in the assignment
of the NH peak at 11.95 ppm for the case of 5g, to the E isomer.
The low energy resonance of the 1,3-thiazolidin-4-one NH is in
accordance with previous results.36 According to our opinion,
intermolecular hydrogen bonds may occur.
Stereochemistry on the C@N imino and C@C exocyclic double
bonds is determined based on NMR spectroscopy. Although syn-
thetic procedures may provide both E and Z isomers of compounds
5a–5j, during recrystallization procedure a pure product of one iso-
mer may occur. This becomes apparent from NMR spectral peaks
and their integration values for almost all compounds (except
compound 5f). More specifically, using DMSO-d6 as solvent, only
one peak was observed for NH at ꢀ12.5 ppm (apart from com-
pound 5f where two NH signals are apparent at 12.71 ppm and
11.95 ppm) resulting in the predominant existence of one isomer.
More specifically, Z configuration of the exocyclic C@C bond, in
the 5-benzylidene derivatives was attributed, since the methine
Aromatic ring
Thiazoyl H
=CH
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmc.2009.10.041.
1.50
1.60
1.70
1.80
1.90
2.00
2.10
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