Thiazolidione derivatives as novel antibiofilm agents: Design, synthesis, biological evaluation, and structure-activity relationships

Article abstract of DOI:10.1016/j.ejmech.2010.12.014

Rational designed novel thiazolidiones were synthesized and evaluated for antibiofilm activity. The active derivatives were not only potent inhibitors of Staphylococcus epidermidis biofilm growth but also efficient antibacterial agents. 3f showed 4-fold higher activity (6.25 μM) in the biofilms dispersal assay and significantly higher antibacterial activity (MIC 3.125 μM) in comparison to the 3-(5-((6- (ethoxycarbonyl)-5-(benzo[1,3]dioxol-5-yl)-3-oxo-7- phenyl- thiazolo[3,2-a]pyrimidin-2(5H)-ylidene)methyl)furan-2-yl)benzoic acid (1).

Full text of DOI:10.1016/j.ejmech.2010.12.014

European Journal of Medicinal Chemistry 46 (2011) 819e824
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Thiazolidione derivatives as novel antibiofilm agents: Design, synthesis, biological
evaluation, and structureeactivity relationships
,
,
,
,
b
,
a
Bin Pan a 1, Renzheng Huang b 1, Likang Zheng a 1, Chen Chen a, Shiqing Han a , Di Qu , Mingli Zhu ,
*
*
Ping Wei a
a College of Biotechnology and Pharmaceutical Engineering, Nanjing University of Technology, Nanjing 210009, China
b Key Laboratory of Medical Molecular Virology of Ministry of Education and Ministry of Pulic Health, Institute of Medical Microbiology and Institutes of Biomedical Sciences,
Shanghai Medical School of Fudan University, Box 228, Shanghai 200032, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Rational designed novel thiazolidiones were synthesized and evaluated for antibiofilm activity. The
active derivatives were not only potent inhibitors of Staphylococcus epidermidis biofilm growth but also
Received 17 August 2010
Received in revised form
5 December 2010
Accepted 10 December 2010
Available online 21 December 2010
efficient antibacterial agents. 3f showed 4-fold higher activity (6.25
mM) in the biofilms dispersal assay
and significantly higher antibacterial activity (MIC 3.125 M) in comparison to the 3-(5-((6- (ethox-
m
ycarbonyl)-5-(benzo[1,3]dioxol-5-yl)-3-oxo-7-phenyl- thiazolo[3,2-a]pyrimidin-2(5H)-ylidene)methyl)
furan-2-yl)benzoic acid (1).
Ó 2010 Elsevier Masson SAS. All rights reserved.
Keywords:
Thiazolidione
Staphylococcus epidermidis
Antibiofilm activity
Inhibitors
1. Introduction
but also efficient antibacterial agent [8]. It elicits effect through
inhibition of the YycG histidine kinase, which is an important
Bacterial biofilms are known to infect patients with in-dwelling
medical devices (IMDs) such as cardiac pacemakers and heart
stents [1e3]. They also possess increased resistance to host
immune responses and antibiotics, often representing a significant
challenge to overcome by the increased morbidity and mortality
rates of numerous biofilm-mediated diseases [4e7]. Recent esti-
mates have attributed biofilm-associated infections as being
responsible for upward of 75% of microbial infections. Given the
breadth of biofilm-mediated infections, there is a significant need
for novel antibiofilm modulators.
control protein of two-component system (TCS) YycG/YycF regu-
lated biofilm formation positively. Its unique mechanism of action
underlies the efficacy and low risk of side effects. However, the
improvement of antibiofilm potency of 1 is still needed and its
structure modification is a challenging task.
We have recently focused on the development of straightfor-
ward chemistry to access analogues of thiazolidione for antibiofilm
screening [9]. In a continued effort, we sought to a rational design
to modify functional groups and to alter scaffold of compound 1 in
generating derivatives to improve the antibiofilm and antibacterial
activities. Additionally, the structureeactivity relationships (SAR) of
the thiazolidione family in the context of antibiofilm activity were
also critical problems which we attempted to probe in this article.
In a first effort to optimize the activity of compound 1 (Scheme
1), C-5 position was evaluated utilizing phenyl and 4-methoxy
phenyl substituents (1aeb). For C-2 position methyl 3-(5-for-
mylfuran-2-yl)benzoate fragment (1ced) and diverse substituted
phenyl groups were introduced (1eej). To further explore features
which might be necessary for antibiofilm activity of compound 1,
we paid attention to phenoxy acetic acid derivatives (PAADs).
PAADs were reported as potent antimycobacterial agents [10].
Following the same rationale of multitarget-directed ligands
As shown in Fig. 1, compound 1 derived from the thiazolidione
class is a pioneering antibiofilm agent which is not only a moderate
inhibitor of Staphylococcus epidermidis (ATCC35984) biofilm growth
Abbreviations: IMDs, in-dwelling medical devices; TCS, two-component system;
PAADs, phenoxy acetic acid derivatives; SAR, structureeactivity relationship;
MTDLs, multitarget-directed ligands; DHPMs, dihydropyrimidine scaffold; PBS,
phosphate buffered saline; TSB, tryptic soy broth; MIC, minimal inhibitory
concentration; CLSI, Clinical and Laboratory Standards Institute.
* Corresponding authors. Tel.: þ86 25 83587334; fax: þ86 25 83587326.
E-mail addresses: hanshiqing@njut.edu.cn (S. Han), dqu@shmu.edu.cn (D. Qu).
These authors contributed equally to this work.
1
0223-5234/$ e see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.12.014

820
B. Pan et al. / European Journal of Medicinal Chemistry 46 (2011) 819e824
R1
R1
N
O
C2H5OOC
N
1e-j
R2
S
COOH
R3
O
N
Functional groups modification
N
R2
O
O
PAADs
O
Pharmacophore overlapping
O
R1
O
R1
N
O
O
C2H5OOC
C2H5OOC
5
C2H5OOC
N
N
N
2
B
A
S
N
S
O
S
O
N
R2
O
COOH
1a-d
1
2a , 2b
COOH
Ring opening
R1
N
R1
N
O
N
O
Chain shortening
S
O
R2
S
R2
N
R1
R1
3c-f
3a, 3b
Fig. 1. Design Strategy.
(MTDLs) [11], we envisioned that merging the antimycobacterial
features of PAADs and antibiofilm features of compound 1 in
a single chemical entity could, hopefully, lead to agents with
multiple antibacterial mechanisms, hence a better potential for
prevention and therapy of biofilm-mediated diseases. Therefore,
we replaced one terminal 3-(5-formylfuran-2-yl)benzoic acid
function group of 1 with a structure-similar 2-phenoxyacetic acid
fragment to afford new compounds (2a, 2b). The structure of PAADs
also inspired that opening the B-ring of compound 1 but keep its
thiazolidione core structure stability would furnish a novel class of
compounds contained more flexible aryl side chains connecting
with N atoms (3aeb). To further explore the SAR, derivatives 3cef
were designed and synthesized by shortening the chain length
between N atoms and phenyl groups.
2. Chemistry
The general synthesis of 1aej was shown in Scheme 1. A simple
one-pot reaction known as Biginelli reaction of phenylaldehyde or
substituted phenylaldehyde, ethyl benzoylacetate and thiourea
gave different dihydropyrimidine scaffolds (DHPMs, 4 ,5 and 6)
[12]. Cyclization of the three DHPMs with chloracetic acid afforded
7, 8 and 9 [13]. An aldol condensation of substituted aromatic
aldehydes with equivalent amount of 7 (8 or 9) was catalysed by
acid or base to give 1aej in good yields [14,15]. As shown in Scheme
2, compounds 2a and 2b were readily prepared in two steps from 1i
and 1j [16e18]. Cyclization reaction, aldol condensation, substitu-
tion reaction and hydrolization were efficiently used to synthesize
3aef, as shown in Scheme 3 [9,15].
R1 CHO
R1
R1
N
R1
O
O
+
C2H5OOC
C2H5OOC
R1=Ph, R2=COOH
C2H5OOC
1a :
1b :
NH
N
S
N
iii
i
ii
R1=4-OCH3-Ph, R2=COOH
S
S
N
H
S
O
N
H2N
NH2
O
R2
1c :
1d :
R1=Ph, R2=COOCH3
+
R =4-OCH -Ph, R =COOCH
3
1
3
2
4 :
R1=4-OCH3-Ph
7 :
R1=4-OCH3-Ph
iv
C2H5OOC
Ph
5 :
6 :
8 :
R1=Ph
O
R1=
O
R1=Ph
O
R =4-OCH -Ph, R =H, R =OH
1e :
R1
N
1
3
2
3
9 : R1=
O
O
C2H5OOC
1f :
R1=4-OCH3-Ph, R2=N(CH3)2, R3=H
R1=4-OCH3-Ph, R2=H, R3=NO2
R1=4-OCH3-Ph, R2=OH, R3=OCH3
N
1g :
S
R3
1h :
1i :
R1=4-OCH3-Ph, R2=OH, R3=H
O
R2
R2=OH, R3=H
R1=
1j :
O
Scheme 1. Reagents and conditions: (i) SnCl2$2H2O, EtOH, reflux; (ii) chloroacetic acid, NaOAc, HAc/Ac2O, reflux; (iii) b-alanine, acetic acid, reflux; (iv) piperidine, absolute alcohol,
60 ꢀC.

B. Pan et al. / European Journal of Medicinal Chemistry 46 (2011) 819e824
821
R1
R1
O
O
C2H5OOC
C2H5OOC
2a :
R1=
R1=
i
ii
N
N
1i, 1j
OCH3
O
S
S
N
N
2b :
COOH
Scheme 2. Reagents and conditions: (i) BrCH2COOCH3, acetone, reflux; (ii) K2CO3, water, methanol, reflux.
O
O
O
COOCH3
3. Results and discussion
several compounds exceeded or maintained the potency of known
biofilms inhibitor thiazolo[3,2-a]pyrimidin 1. In this context, some
points were noteworthy: (1) B-ring opening analogs of thiazolo[3,2-
a]pyrimidin 1 maintained the antibiofilm potency, (2) substituent
phenyl moieties which connected with N atoms obviously influ-
enced antibiofilm activity, (3) the large-sized carboxylic acid moie-
ties such as 4-(5-formylfuran-2-yl)benzoic acid fragment which
replaced nitrophenyl, hydroxyphenyl or dimethylaminophenyl at C-
2 position obtained excellent activity. Furthermore, the most
promising results were observed for compound 3f, with potent
Among tested compounds (Table 1),1aej did not yield any major
improvements. Only C-5 position 4-methoxy phenyl derivative 1b
displayed comparable antibiofilm and antibacterial activities to
compound 1. Nitrophenyl, hydroxyphenyl, dimethylaminophenyl
and methyl 3-(5-formylfuran-2-yl) benzoate fragment introduced
intoC-2 position led bioactivity disappeared (>200
2a and 2b were both less potent agents than compound 1, with
antibiofilm concentrations of 50 M and MIC values of 25 M,
mM). Bycontrast,
m
m
respectively. These data indicated that 3-(5-formylfuran-2-yl)ben-
zoic acid fragment might be a more potent moiety than 2-phe-
noxyacetic acid fragment. The rigidity of the large moiety, probably,
was one of the factors led the enhancement of activity.
Interestingly, the ring-opening derivative 3a was a more potent
antibiofilm and antibacterial agent than 2a, with antibiofilm
antibiofilm activity (6.25
mM) and high antibacterial activity
(MIC ¼ 3.125 M). In compare with 1, the chemical structure of 3f
m
was simpler and its activity was improved in 4-fold. These findings
provide important pharmacophore information for the exploration
of drug-like antibiofilm agents and further studies are on going in
our laboratories.
concentration 25
mM and MIC 12.5 mM, respectively. Comparing
with 1b, the corresponding ring-opening compound 3b was a less
potent agent. The activity data of 3a and 3b showed that B-ring
opening analogs of thiazolo[3,2-a]pyrimidin 1 maintained the
antibiofilm and antibacterial potency. Surprisingly, the chain-
shortening analogs 3cef (derivatives of 3b) showed excellent
antibiofilm activities among the tested compounds. In comparison
with compound 3b, the antibiofilm concentration and MIC of 3c
were improved in 8-fold (showing 6.25 mM and 6.25 mM, respec-
tively), and this improvement cued that the activity increased
when substituted phenyl moieties linked with N atoms directly. The
5. Experimental
All reagents and solvents were commercially available and used
without further purification. Melting points were determined on an
electrothermal digital apparatus model WRS-1B (Shanghai, China),
without correction. Infrared spectra were recorded on an Avatar
360 ESP spectrometer (Nicolet), using potassium bromide (1% w/w)
disk scanning from 500 to 4000 cmꢁ1. A Bruker AM-500 MHz
instrument (Bruker) was used to acquire 1H NMR and 13C NMR
spectra with TMS as internal standard. Chloroform-D and DMSO-d6
were used as solvents. Low resolution mass spectra (LRMS) were
recorded on an HP-5973 instrument (HP) and High resolution mass
spectra (HRMS) were recorded on FTMS-7.0 instrument (Bruker) or
IonSpec 4.7T instrument (Varian).
most potent inhibitor 3f showed a 4-fold higher activity (6.25
in the ATCC35984 biofilms dispersal assay and significantly higher
M) in comparison to compound
mM)
antibacterial activity (MIC ¼ 3.125
m
1. It was also notable that replacing the 3-benzoic acid substituent
(3e) with 4-benzoic acid substituent (3f) resulted in a 4-fold
The purity of the final compounds was determined by HPLC on
a DIONEX-P680 instrument (DIONEX) at 254 nm and all final
compounds have a purity of >95%.
improvement in its antibiofilm concentration (6.25
mM) and MIC
(3.125 M). All the results showed that fragments connected with
m
C-2 position and the substituent phenyl moieties connected with N
atoms might be two decisive factors influenced the activity.
5.1. General procedure for preparation of compounds (1aed)
4. Conclusion
The corresponding aldehyde (1 equiv), b-alanine (2 equiv), and 7
(or 8) (2 equiv) were heated at 100 ꢀC for 1 h in glacial acetic acid.
Upon completion of the reaction, the mixture was cooled, the
reaction was quenched with water, and the precipitate was filtered
A series of thiazolidione derivatives had been designed, synthe-
sized and investigated as antibiofilm agents to the S. epidermidis,
O
O
N
S
R1
R1
N
R1
N
H
N
H
N
R1
i
iii
ii
iv
v
N
N
3a : R1=4-OCH3-Ph-CH2-
H2N R1
R1
R1
R1
R1
O
S
S
S
N
R1=4-OCH3-Ph-CH2-
R1=2-CH3-Ph-
10 : R1=4-OCH3-Ph-CH2-
14
11 :
R1=2-CH3-Ph-
vi
OH
O
R1=4-CH3-Ph-
R1=4-OCH3-Ph-
12 :
O
R1=4-CH3-Ph-
R1=4-OCH3-Ph-
COOH
R1
N
13 :
N
R1
3b :
R1=4-OCH3-Ph-CH2-, R2=3-COOH
3c : R1=4-OCH3-Ph-, R2=3-COOH
S
O
R2
3d :
R1=4-CH3-Ph-, R2=3-COOH
3e :
R1=2-CH3-Ph-, R2=3-COOH
3f
:
R1=2-CH3-Ph-, R2=4-COOH
Scheme 3. Reagents and conditions: (i) CS2, Lac.sulfur, triethanolamine, water, reflux; (ii) chloroacetic acid, NaOAc, absolute alcohol, reflux; (iii) piperidine, absolute alcohol, 60 ꢀC;
(iv) BrCH2COOCH3, acetone, reflux; (v) K2CO3, water, methanol, reflux; (vi) -alanine, acetic acid, reflux.
b

822
B. Pan et al. / European Journal of Medicinal Chemistry 46 (2011) 819e824
([M þ H]þ), 613.4([M þ Na]þ); HRMS calcd for C34H27N2O6S [M þ H]þ
Table 1
Antibiofilm and anti-bacteria activity of thiazolidione derivatives.
591.1590, found 591.1586.
Compound
Antibiofilm concentration(
m
M)
MIC(
>200a
6.25
>200
>200
>200
>200
>200
>200
>200
25
m
M)
5.1.4. Ethyl 2-((5-(3-(methoxycarbonyl)phenyl)furan-2-yl)methylene)-
5-(4-methoxyphenyl)-3- oxo-7-phenyl-thiazolo[3,2-a]pyrimidine-6-
carboxylate (1d)
Prepared from compound 7 and methyl 3-(5-formylfuran-2-yl)
benzoate in 90% overall yield and obtained as a yellow solid. mp:
248e249 ꢀC; IR(KBr), v(cmꢁ1): 3030,1715,1548,1244,1175,1031; 1H
1a
1b
1c
1d
1e
1f
1g
1h
1i
2a
2b
3a
3b
3c
3d
3e
3f
>200
25
>200
>200
>200
>200
>200
>200
>200
50
50
25
100
6.25
12.5
25
6.25
25
NMR (DMSO-d6)
d
8.39 (s, 1H), 8.10 (d, J ¼ 8.2 Hz, 1H), 7.96
(t, J ¼ 8.1 Hz, 1H), 7.72e7.67 (m, 2H), 7.46e7.38 (m, 6H), 7.33
(d, J ¼ 8.8 Hz, 2H), 7.28 (d, J ¼ 3.7 Hz,1H), 6.94 (d, J ¼ 8.8 Hz, 2H), 6.11
(d, J ¼ 7.1 Hz, 1H), 3.90 (s, 3H), 3.85e3.79 (m, 2H), 3.73 (s, 3H), 0.81
25
12.5
50
6.25
6.25
12.5
(t, J ¼ 7.1 Hz, 3H); 13C NMR (DMSO-d6)
d
165.6, 165.4, 163.9, 159.4,
155.6,149.2,138.5,131.9,130.6,129.8,129.1,128.8,128.5,128.2,127.6,
124.6,121.4,118.4,116.9,114.1,110.7,110.1, 60.1, 55.1, 52.4,13.2; HRMS
calcd for C35H29N2O7S [M þ H]þ 621.1697, found 621.1690.
3.125
6.25
1
5.2. General procedure for preparation of compounds (1eej)
a
The initiate dilution was 200 mM.
A mixture of corresponding benzaldehyde (3 mmol), 7 (or 9)
(3 mmol) in 15 mL ethanol in the presence of piperidine (5 mmol)
was refluxed for 6 h. The mixture was cooled to room temperature
and extracted with EtOAc. The organic layer was washed with
brine, dried by anhydrous Na2SO4, and concentrated under reduced
pressure. The crude compound was purified by column chroma-
tography (stationary phase, silica; mobile phase, petroleum ether/
ethyl acetate 8/1).
off. The solid products were filtered and recrystallized in methanol
to obtain pure compound.
5.1.1. 3-(5-((6-(Ethoxycarbonyl)-3-oxo-5,7-diphenyl-thiazolo[3,2-a]
pyrimidin-2(5H)-ylidene)methyl) furan-2-yl)benzoic acid (1a)
Prepared from compound 8 and 3-(5-formylfuran-2-yl)benzoic
acid in 93% overall yield and obtained as a yellow solid. mp:
284e285 ꢀC; IR(KBr): v(cmꢁ1): 3387, 3010, 1698, 1609, 1549, 1329,
5.2.1. Ethyl 2-(3-hydroxybenzylidene)-5-(4-methoxyphenyl)-3-oxo-7-
phenyl-thiazolo [3,2-a] pyrimidine-6-carboxylate (1e)
1182, 1079; 1H NMR (DMSO-d6)
d
¼ 8.38(s, 1H, CH), 8.09e8.07
(d, J ¼ 6.0 Hz, 1H), 7.97e7.95 (d, J ¼ 6.0 Hz, 1H), 7.69e7.41 (m, 14H),
6.16 (s, 1H), 3.86e3.79(q, J ¼ 6.9 Hz, 2H), 0.82e0.77 (t, J ¼ 6.0 Hz,
Prepared from compound 7 and 3-hydroxybenzaldehyde in 90%
overall yield and obtained as a yellow solid. mp: 187e188 ꢀC;
IR(KBr), v(cmꢁ1): 3403, 2992, 1700, 1609, 1541, 1327, 1244, 1175,
3H); 13C NMR (DMSO-d6)
d 166.7, 165.4, 163.9, 156.0, 155.9, 149.6,
149.1, 139.9, 138.5, 131.8, 129.5, 128.9, 128.8, 128.6, 128.2, 127.6,
127.3, 124.7, 121.6, 118.6, 116.6, 110.5, 109.9, 60.1, 55.7, 13.2; ESI-
MS m/z 575.2 [M ꢁ H]þ; HRMS calcd for C33H25N2O6S [M þ H]þ
577.1433, found 577.1430.
1061; 1H NMR (DMSO-d6)
d 9.86 (s, 1H), 7.71 (s, 1H), 7.41
(d, J ¼ 3.2 Hz, 5H), 7.33 (d, J ¼ 8.8 Hz, 3H), 7.05 (d, J ¼ 8.4 Hz,1H), 6.98
(d, J ¼ 1.8 Hz,1H), 6.94 (d, J ¼ 8.8 Hz, 2H), 6.89 (dd, J ¼ 7.8, 2.0 Hz,1H),
6.11 (s,1H), 3.85e3.78 (m, 2H), 3.73 (s, 3H), 0.79 (t, J ¼ 7.1 Hz, 3H); 13C
NMR (DMSO-d6) d 165.4,164.2,159.4,157.8,154.9,149.1,138.4,133.9,
5.1.2. 3-(5-((6-(Ethoxycarbonyl)-5-(4-methoxyphenyl)-3-oxo-7-
phenyl- thiazolo[3,2-a]pyrimidin-2(5H)-ylidene)methyl)furan-2-yl)
benzoic acid (1b)
133.0, 131.7, 130.4, 128.9, 128.6, 128.1, 127.6, 121.1, 119.6, 118.0, 115.9,
114.1, 110.4, 60.1, 55.1, 13.2; HRMS calcd for C29H24N2O5S (Mþ)
512.1404, found 512.1406.
Prepared from compound 7 and 3-(5-formylfuran-2-yl)ben-
zoic acid in 92% overall yield and obtained as a yellow solid. mp:
279e280 ꢀC; IR(KBr), v(cmꢁ1): 3371, 3053, 1708, 1615, 1547, 1330,
5.2.2. Ethyl 2-(4-(dimethylamino)benzylidene)-5-(4-methoxyphenyl)-
3-oxo-7-phenyl-thiazolo [3,2-a] pyrimidine-6-carboxylate (1f)
Prepared from compound 7 and 4-(dimethylamino)benzalde-
hyde in 89% overall yield and obtained as a yellow solid. mp:
202e203 ꢀC; IR(KBr), v(cmꢁ1): 2984, 1707, 1571, 1533, 1236, 1168,
1181, 1069; 1H NMR (DMSO-d6)
d
¼ 8.34 (s, 1H), 8.05e8.03
(d, J ¼ 6.0 Hz, 1H), 7.93e7.91 (d, J ¼ 6.0 Hz, 1H), 7.64e6.92 (m,
13H), 6.06 (s, 1H), 3.80e3.78 (q, J ¼ 6.0 Hz, 2H), 3.69 (s, 3H),
0.79e0.75 (t, J ¼ 6.0 Hz, 3H); 13C NMR (DMSO-d6)
d
166.6, 165.4,
1053; 1H NMR (DMSO-d6)
d
7.68 (s, 1H), 7.44 (d, J ¼ 9.0 Hz, 1H), 7.39
163.9, 159.4, 155.9, 155.6, 149.4, 149.1, 138.6, 131.9, 129.5, 128.9,
128.5, 128.2, 127.6, 124.7, 121.5, 118.5, 116.7, 114.1, 110.5, 110.1,
60.1, 55.1, 13.3; HRMS calcd for C34H27N2O7S [M þ H]þ 607.1529,
found 607.1533.
(s, 5H), 7.32 (d, J ¼ 8.7 Hz, 2H), 6.93 (d, J ¼ 8.7 Hz, 2H), 6.82
(d, J ¼ 9.1 Hz, 2H), 6.11 (s, 1H), 3.86e3.77 (m, 2H), 3.73 (s, 3H), 3.02
(s, 6H), 0.80 (t, J ¼ 7.1 Hz, 3H); 13C NMR (DMSO-d6)
d 165.5, 164.5,
159.3,155.8,151.6,150.1,138.8, 134.1,132.2,128.7,128.5,128.2, 127.6,
119.5, 114.1, 112.0, 111.6, 111.2, 109.4, 59.9, 55.0, 54.7, 13.2; HRMS
calcd for C31H29N3O4S (Mþ) 539.1880, found 539.1879.
5.1.3. Ethyl 2-((5-(3-(methoxycarbonyl)phenyl)furan-2-yl)methylene)-
3-oxo-5,7-diphenyl- thiazolo[3,2-a]pyrimidine-6-carboxylate (1c)
Prepared from compound 8 and 3-(5-formylfuran-2-yl)benzoate
in 90% overall yield and obtained as a yellow solid. mp: 262e263 ꢀC;
IR(KBr), v(cmꢁ1): 3023, 1723, 1639, 1578, 1236, 1160, 933; 1H NMR
5.2.3. Ethyl 2-(3-nitrobenzylidene)-5-(4-methoxyphenyl)-3-oxo-7-
phenyl- thiazolo[3,2-a]pyrimidine-6-carboxylate (1g)
Prepared from compound 7 and 3-nitrobenzaldehyde in 90%
overall yield and obtained as a yellow solid. mp: 144e145 ꢀC; IR
(KBr), v(cmꢁ1): 2984, 1693, 1541, 1335, 1244, 1191, 1138, 1061; 1H
(DMSO-d6)
d
8.39 (s, 1H), 8.11 (d, J ¼ 7.9 Hz, 1H), 7.97 (d, J ¼ 7.8 Hz,
1H), 7.73e7.68 (m, 2H), 7.53e7.20 (m, 12H), 6.16 (d, J ¼ 7.9 Hz, 1H),
3.90 (s, 3H), 3.83 (q, J ¼ 7.0 Hz, 2H), 0.80 (t, J ¼ 7.1 Hz, 3H); 13C NMR
NMR (DMSO-d6)
d 7.98 (s, 1H), 7.45e7.37 (m, 6H), 7.36e7.32
(DMSO-d6)
d
165.7, 165.4, 163.9, 155.8, 155.7, 149.5, 149.2, 139.8,
(m, 2H), 7.32e7.26 (m, 2H), 7.00e6.92 (m, 2H), 6.13 (s, 1H),
138.5,130.6,129.9,129.2,128.9,128.7,128.6,127.7,127.4,124.6,121.6,
118.6, 116.8, 110.8, 110.0, 60.1, 55.7, 52.4, 13.2; ESI-MS m/z 591.2
3.86e3.79 (m, 2H), 3.74 (s, 3H), 0.80 (t, J ¼ 7.1 Hz, 3H); 13C NMR
(DMSO-d6)
d 165.4, 163.8, 159.5, 154.2, 148.7, 148.2, 138.2, 135.1,

B. Pan et al. / European Journal of Medicinal Chemistry 46 (2011) 819e824
823
134.5, 131.4, 130.9, 130.3, 129.4, 128.2, 127.9, 124.5, 114.0, 110.8, 60.2,
55.3, 55.1, 13.2; HRMS calcd for C31H29N3O4S (Mþ) 541.1307, found
541.1308.
(DMSO-d6) d 165.5, 164.4, 163.1, 160.1, 159.4, 155.2, 149.52, 138.6,
132.9, 131.9,131.5,128.9, 128.6,128.2,127.6,125.4, 116.6,115.5,115.0,
114.1, 110.1, 60.1, 55.1, 13.3; ESI-MS m/z 571.1([M þ H]þ), 593.1
([M þ Na]þ); HRMS calcd for C31H27N2O7S [M þ H]þ 571.1539,
found 571.1533.
5.2.4. Ethyl 2-(4-hydroxy-3-methoxybenzylidene)-5-(4-methoxyph
enyl)-3-oxo-7-phenyl- thiazolo [3,2-a] pyrimidine-6-carboxylate (1h)
Prepared from compound 7 and 4-hydroxy-3-methoxybenzalde
hyde in 85% overall yield and obtained as a yellow solid. mp:
153e154 ꢀC;IR(KBr),v(cmꢁ1):3418,2984,1707,1541,1502,1236,1175,
5.3.2. 2-(4-((5-(Benzo[d][1,3]dioxol-5-yl)-6-(ethoxycarbonyl)-3-oxo
-7-phenyl-thiazolo[3,2-a]pyrimidin-2(5H)-ylidene)methyl)phenoxy)
acetic acid (2b)
1061; 1H NMR (DMSO-d6)
d
10.00 (s,1H), 7.73 (s,1H), 7.41 (s, 5H), 7.33
Prepared from compound 1j in overall yield 65% and obtained as
(d, J ¼ 8.8 Hz, 2H), 7.17 (d, J ¼ 2.0 Hz,1H), 7.09 (dd,1H), 6.98e6.91 (m,
a yellow solid. 1H NMR (300 MHz, DMSO-d6)
d
¼ 7.79(s, 1H),
3H), 6.12 (s,1H), 3.86e3.78(m, 5H), 3.73 (s, 3H), 0.80(t, J ¼ 7.1 Hz, 3H);
7.59e6.89(m, 12H), 6.08(s, 1H), 6.02(s, 2H), 4.78(s, 2H), 3.86e3.79
(q, J ¼ 6.9 Hz, 2H), 0.82e0.78(t, J ¼ 6.0 Hz, 3H); ESI-MS m/z 585.4
([M þ H]þ),583.5([M ꢁ H]þ).
13C NMR (DMSO-d6)
d 165.4, 164.4, 159.4, 155.4, 149.8, 149.5, 148.0,
138.5, 133.7, 131.9, 128.7, 128.5, 128.2, 127.6, 124.2, 116.2, 115.5, 114.0,
109.9, 60.0, 55.6, 55.0, 54.9, 13.2; ESI-MS m/z 543.1([M þ H]þ), 565.3
([M þ Na]þ); HRMS calcd for C33H27N2O6S [M þ H]þ 543.1569, found
543.1584.
5.4. 5-(4-Hydroxybenzylidene)-3-(4-methoxybenzyl)-2-(4-methox
ybenzylimino)thiazolidin-4-one (14)
5.2.5. Ethyl 2-(4-hydroxybenzylidene)-5-(4-methoxyphenyl)-3-oxo
-7-phenyl- thiazolo[3,2-a]pyrimidine-6-carboxylate (1i)
Prepared from compound 7 and 4-hydroxybenzaldehyde in 87%
overall yield and obtained as a yellow solid. mp: 150e151 ꢀC; IR
(KBr), v(cmꢁ1): 3418, 2904, 1700, 1548, 1510, 1235, 1160, 1053; 1H
A mixture of 4-hydroxybenzaldehyde (3 mmol), compound 10
(3mmol)in15 mLethanolinthepresenceofpiperidine(5mmol)was
refluxed for 6 h. Upon completion of the reaction, the reaction
mixture was extracted with EtOAc. The organic layer was washed
with brine, dried by anhydrous Na2SO4, and concentrated under
reduced pressure. The crude compound was purified by silica gel
column chromatography using petroleum ether/ethyl acetate (4:1 v/
v) as mobile phase to obtain pure compound 14 as a yellow solid:
NMR (DMSO-d6)
d 10.35 (s, 1H), 7.72 (s, 1H), 7.51e7.46 (m, 2H),
7.42e7.38 (m, 5H), 7.35e7.29 (m, 2H), 6.97e6.89 (m, 4H), 6.11
(s, 1H), 3.86e3.77 (m, 2H), 3.72 (s, 5H), 0.80 (t, J ¼ 7.1 Hz, 3H); 13C
NMR (DMSO-d6)
d
165.4, 164.5, 160.2, 159.4, 155.4, 149.6, 138.6,
yield, 95%; 1H NMR (CDCl3)
d
¼ 7.69 (s, 1H), 7.44e6.80 (m, 12H), 5.05
133.4, 132.4, 132.0, 128.8, 128.5, 128.2, 127.6, 123.7, 116.4, 115.8,
115.2, 114.1, 110.0, 60.0, 55.0, 54.9, 13.2; ESI-MS m/z 513.1
([M þ H]þ), 535.0([M þ Na]þ); HRMS calcd for C29H25N2O5S
[M þ H]þ 513.1487, found 513.1478.
(s, 2H), 4.58 (s, 2H), 3.81 (s, 3H), 3.77 (s, 3H); ESI-MS m/z 461.1
([M þ H]þ), 483.2([M þ Na]þ).
5.5. 2-(4-((3-(4-Methoxybenzyl)-2-(4-methoxybenzylimino)-4-oxo
thiazolidin-5-ylidene)methyl) phenoxy)acetic acid (3a)
5.2.6. Ethyl 2-(4-hydroxybenzylidene)-5-(benzo[d][1,3]dioxol-5-yl)-
3-oxo-7-phenyl-thiazolo [3,2-a] pyrimidine-6-carboxylate (1j)
Prepared from compound 9 and 4-hydroxybenzaldehyde in 85%
overall yield and obtained as a yellow solid. mp: 160e161 ꢀC; 1H
A mixture of compound 14 (1 mmol), methyl bromoacetate
(1.5 mmol) and potassium carbonate (1.5 mmol) in acetone (15 ml)
was refluxed for 6 h. The solvent was evaporated under reduced
pressure and the solid residue was purified by silica gel column
chromatography using petroleum ether/ethyl acetate (4:1 v/v) as
mobile phase. The purified solid (1 mmol) and potassium carbonate
(2 mmol) in the medium of 25 ml methanol and 15 ml water was
refluxed for 12 h. The reaction mixture was then filtered and the
filtrate was acidified with HCl until solid precipitated. The solid
products were filtered and recrystallized in methanol to obtain pure
compound as a yellow solid: yield, 70%; mp: 196e197 ꢀC; IR(KBr), v
(cmꢁ1): 3327, 2992,1746,1685,1647,1510,1244; 1H NMR (DMSO-d6)
NMR (300 MHz, CDCl3)
d
¼ 8.02 (s, 1H), 7.38e7.00 (m, 12H), 6.23
(s, 1H), 5.92 (s, 2H), 3.94e3.87 (q, J ¼ 6.9 Hz, 2H), 0.90e0.86
(t, J ¼ 6.0 Hz, 3H); 13C NMR (CDCl3):
d 166.3, 165.7, 158.9, 157.2,
151.5, 148.3, 148.2, 139.0, 134.5, 133.7, 132.6, 129.1, 128.4, 128.2,
125.5, 122.2, 116.7, 110.3, 108.6, 101.5, 61.0, 56.0, 13.7; ESI-MS m/z
527.1([M þ H]þ), 549.2([M þ Na]þ).
5.3. General procedure for preparation of compounds (2a, 2b)
A mixture of compound 1i or 1j (1 mmol), methyl bromoacetate
(1.5 mmol) and potassium carbonate (1.5 mmol) in acetone (15 ml)
was refluxed for 4 h. The solvent was evaporated under reduced
pressure and the solid residue was purified by silica gel column
chromatography using petroleum ether/ethyl acetate (4:1 v/v) as
mobile phase. The purified solid (1 mmol) and potassium carbonate
(1 mmol) in the medium of 30 ml methanol and 20 ml water was
refluxed for 12 h. The reaction mixture was then filtered and the
filtrate was acidified with HCl until solid precipitated. The solid
products were filtered and recrystallized in methanol to obtain
pure compound 2a or 2b.
d
¼ 7.70 (s, 1H), 7.62e6.85 (m, 12H), 4.92 (s, 2H), 4.76 (s, 2H), 4.54 (s,
2H), 3.73 (s, 3H), 3.71 (s, 3H); 13C NMR (DMSO-d6)
d
169.7, 165.8,
159.0,158.6,158.1,148.0,131.7,130.9,129.4,129.3,128.4,126.3,118.3,
115.3,113.7,113.7, 64.5, 55.0, 54.9, 45.0; ESI-MS m/z 519.0([M þ H]þ);
HRMS calcd for C28H27N2O6S [M þ H]þ 519.1590, found 519.1591.
5.6. The synthesis of 3-(5-((3-(4-methoxybenzyl)-2-(4-methoxyben
zylimino)-4-oxothiazolidin-5-ylidene)methyl) furan-2-yl) benzoic
acid (3b) illustrates the general sequence used to prepare compounds
3bef
5.6.1. 3-(5-((3-(4-Methoxybenzyl)-2-(4-methoxybenzylimino)-4-
5.3.1. 2-(4-((6-(Ethoxycarbonyl)-5-(4-methoxyphenyl)-3-oxo-7-
phenyl-thiazolo [3,2-a] pyrimidin-2(5H)-ylidene)methyl)phenoxy)
acetic acid (2a)
oxothiazolidin-5-ylidene)methyl) furan-2-yl)benzoic acid (3b)
A mixture of methyl 3-(5-formylfuran-2-yl)benzoate (3 mmol), b-
alanine (3 mmol), and compound 10 (3 mmol)were heated at 100 ꢀC
for 1 h in acetic acid (10 mL). Upon completion of the reaction, the
mixture was cooled, the reaction was quenched with water (20 mL),
and the precipitate was filtered off. The solid was washed with water
and methanol and then finally dried with diethyl ether to give the
desired compound as a yellow solid: yield, 92%; mp: 245e246 ꢀC; IR
Prepared from compound 1i in overall yield 71% and obtained as
a yellow solid. mp: 185e186 ꢀC; IR(KBr), v(cmꢁ1): 3334, 2984, 1715,
1548, 1502, 1244,1175, 1069; 1H NMR (300 MHz, DMSO-d6)
d
¼ 7.73
(s, 1H), 7.62e6.93(m, 13H), 6.20(s, 1H), 4.85(s, 1H), 3.87e3.84
(q, J ¼ 3.0 Hz, 2H), 3.76(s, 3H), 0.86e0.82(t, J ¼ 6.0 Hz, 3H); 13C NMR

824
B. Pan et al. / European Journal of Medicinal Chemistry 46 (2011) 819e824
(KBr), v(cmꢁ1): 3342, 2993, 1693, 1638, 1502, 1252; 1H NMR (DMSO-
d6)
¼ 8.45 (s, 1H), 8.05e8.03 (d, J ¼ 6.0 Hz, 1H), 7.95e7.93 (d,
J ¼ 6.0 Hz, 1H), 7.63e6.87 (m, 12H), 4.92 (s, 2H), 4.62 (s, 2H), 3.72 (s,
6H); 13C NMR (DMSO-d6)
166.8, 165.3, 158.6, 158.1, 155.0, 149.5,
one of the most frequent causes of infections in connection with
surgically implanted medical devices [19,20]. In the antibiofilm
concentration experiment, an overnight culture of S. epidermidis
strain ATCC35984 was diluted 1:200 in TSB containing 0.25%
d
d
148.8, 131.7, 131.0, 129.5, 129.3, 129.1, 128.6, 128.4, 128.0, 124.6, 119.8,
118.5,115.7,113.7,110.0, 55.0, 54.6, 44.8; ESI-MS m/z 555.1([M þ H]þ);
HRMS calcd for C31H27N2O6S [M þ H]þ 555.1590, found 555.1585.
glucose, and then 200 ml bacterial suspension was added into the
wells of sterile 96-well polystyrene microtiter plates (Falcon)
incubated at 37 ꢀC for 6 h. The plates with young biofilm were
washed gently four times with sterile PBS before adding fresh TSB
containing the various derivatives at gradient dilution, and incu-
bated at 37 ꢀC for 18 h. The plates were washed again four times
with PBS, and 99% methanol was added for 15 min. After air-dried,
the biofilm was dyed by crystal violet. A well with young biofilm
washed by sterile PBS then dried was served as negative control;
this value was subtracted from the experimental readings. Another
well with young biofilm added sterile TSB was served as positive
control. Each assay was performed in triplicate. The experiment
was repeated three times. MIC assays for the antibacterial activities
of the compounds were performed according to the broth micro-
dilution (in tubes) method of the Clinical and Laboratory Standards
Institute (CLSI) of America [21].
5.6.2. 3-(5-((3-(4-Methoxyphenyl)-2-(4-methoxyphenylimino)
-4-oxothiazolidin-5-ylidene)methyl) furan-2-yl)benzoic acid (3c)
Prepared from methyl 3-(5-formylfuran-2-yl)benzoate and com
pound 11 and obtained as a yellow solid: yield, 90%; mp:
219e220 ꢀC; IR(KBr), v(cmꢁ1): 3302, 2984, 1700, 1616, 1510, 1244;
1H NMR (DMSO-d6)
d
¼ 8.30(s, 1H), 7.94e7.92(d, J ¼ 6.0 Hz, 1H),
7.89e7.87(d, J ¼ 6.0 Hz, 1H), 7.64e6.97(m, 12H), 3.82(s, 3H), 3.79
(s, 3H); 13C NMR (DMSO-d6)
166.7, 165.3, 159.1, 156.5, 155.0, 150.8,
d
149.7, 140.7, 131.8, 129.6, 129.2, 127.7, 124.5, 122.2, 119.5, 119.0, 116.2,
114.5, 114.1, 110.2, 55.4, 55.3; ESI-MS m/z 525.3([M ꢁ H]þ); HRMS
calcd for C29H23N2O6S [M þ H]þ 527.1277, found 527.1275.
5.6.3. 3-(5-((4-Oxo-3-(4-methylphenyl)-2-(4-methylphenylimino)
thiazolidin-5-ylidene)methyl) furan-2-yl)benzoic acid (3d)
Acknowledgements
Prepared from methyl 3-(5-formylfuran-2-yl)benzoate and com
pound 12 and obtained as a yellow solid: yield, 93%, mp: 259e260 ꢀC;
IR(KBr), v(cmꢁ1): 3395, 3038, 1715, 1624, 1510, 1373, 1168; 1H NMR
We would like to thank Professor Guoqiang Lin, Shanghai
Institute of Organic Chemistry, Chinese Academy of Sciences, for
his invaluable academic advice. This work was supported by
National Natural Science Foundation of China Grant No. 21072095,
Grant No. 20942006 (S-Q. Han), Grant No. 30800036 (D. Qu) and
Shanghai Municipal Committee of Science and Technology
(08JC1401600).
(DMSO-d6)
7.87e7.84(d, J¼ 6.0 Hz,1H), 7.64e6.91(m,12H), 2.38(s, 3H), 2.34(s, 3H);
13C NMR (DMSO-d6)
166.7,165.2,155.1,151.0,149.7,145.2,138.1,133.9,
132.4, 131.8, 129.7, 129.3, 128.2, 127.7, 124.5, 120.8, 119.6, 118.9, 116.3,
110.3, 20.7, 20.5; ESI-MS m/z 493.3([M ꢁ H]þ); HRMS calcd for
C29H23N2O4S [M þ H]þ 495.1379, found 495.1382.
d
¼ 13.18(s, 1H), 8.27(s, 1H), 7.94e7.92(d, J ¼ 6.0 Hz, 1H),
d
5.6.4. 3-(5-((4-Oxo-3-(2-methylphenyl)-2-(2-methylphenylimino)
thiazolidin-5-ylidene)methyl) furan-2-yl)benzoic acid (3e)
References
Prepared from methyl 3-(5-formylfuran-2-yl)benzoate and como-
pund 13 and obtained as a yellow solid: yield, 90%, mp: 262e263 ꢀC; IR
(KBr), v(cmꢁ1): 3380, 2984, 1700, 1639, 1541, 1373, 1161; 1H NMR
[1] R. Darouiche, Clin. Infect. Dis. 33 (2001) 1567e1572.
[2] R. Darouiche, I. Raad, S. Heard, J. Thornby, O. Wenker, A. Gabrielli, J. Berg,
N. Khardori, H. Hanna, R. Hachem, N. Engl. J. Med. 340 (1999) 1e8.
[3] R. Donlan, J. Costerton, Clin. Microbiol. Rev. 15 (2002) 167e193.
[4] J. Costerton, P. Stewart, E. Greenberg, Science 284 (1999) 1318e1322.
[5] T. Rasmussen, M. Givskov, Int. J. Med. Microbiol. 296 (2006) 149e161.
[6] J. Costerton, K. Cheng, G. Geesey, T. Ladd, J. Nickel, M. Dasgupta, T. Marrie,
Annu. Rev. Microbiol. 41 (1987) 435e464.
[7] P. Stoodley, K. Sauer, D. Davies, J. Costerton, Annu. Rev. Microbiol. 56 (2002)
187e209.
[8] Z. Qin, J. Zhang, B. Xu, L. Chen, Y. Wu, X. Yang, X. Shen, S. Molin, A. Danchin,
H. Jiang, D. Qu, BMC Microbiol. 6 (2006) 96e114.
(DMSO-d6)
d
¼ 8.23 (s, 1H), 7.93e7.91 (d, J ¼ 6.0 Hz, 1H), 7.83e7.81 (d,
J ¼ 6.0 Hz, 1H), 7.69e6.96 (m, 12H), 2.25 (s, 3H), 2.05 (s, 3H); 13C NMR
(DMSO-d6) d 166.7, 165.0, 155.2, 150.2, 149.6, 146.4, 136.0, 134.3, 131.8,
130.6, 129.2, 127.7, 126.9, 126.7, 124.8, 124.5, 119.9, 119.6, 118.7, 116.7,
110.3, 17.3, 17.0; ESI-MS m/z 495.0([M þ H]þ),517.1([M þ Na]þ); HRMS
calcd for C29H23N2O4S [M þ H]þ 495.1372, found 495.1373.
[9] B. Pan, R.-Z. Huang, S.-Q. Han, D. Qu, M.-L. Zhu, P. Wei, H.-J. Ying, Bioorg. Med.
Chem. Lett. 20 (2010) 2461e2464.
[10] M. Ali, M. Shaharyar, Bioorg. Med. Chem. 15 (2007) 1896e1902.
[11] A. Cavalli, M. Bolognesi, A. Minarini, M. Rosini, V. Tumiatti, M. Recanatini,
C. Melchiorre, J. Med. Chem. 51 (2008) 347e372.
[12] M. Ashok, B. Holla, N. Kumari, Eur. J. Med. Chem. 42 (2007) 380e385.
[13] A. Rashad, H. Sayed, A. Shamroukh, H. Awad, Phosphorus, Sulfur, Silicon Relat.
Elem. 180 (2005) 2767e2777.
[14] R. Ottana, R. Maccari, R. Ciurleo, M. Vigorita, A. Panico, V. Cardile, F. Garufi,
S. Ronsisvalle, Bioorg. Med. Chem. 15 (2007) 7618e7625.
[15] V. Pomel, J. Klicic, D. Covini, D. Church, J. Shaw, K. Roulin, F. Burgat-Charvillon,
D. Valognes, M. Camps, C. Chabert, J. Med. Chem. 49 (2006) 3857e3871.
[16] R. Maccari, R. Ottana, R. Ciurleo, M. Vigorita, D. Rakowitz, T. Steindl, T. Langer,
Bioorg. Med. Chem. Lett. 17 (2007) 3886e3893.
[17] R. Maccari, P. Paoli, R. Ottana, M. Jacomelli, R. Ciurleo, G. Manao, T. Steindl,
T. Langer, M. Vigorita, G. Camici, Bioorg. Med. Chem. 15 (2007) 5137e5149.
[18] S. Wang, Y. Ye, Z. Zhang, R. Tan, Ultrason. Sonochem. 13 (2006) 28e31.
[19] M. Rupp, G. Archer, Clin. Infect. Dis. (1994) 231e243.
5.6.5. 4-(5-((4-Oxo-3-(2-methylphenyl)-2-(2-methylphenylimino)
thiazolidin-5-ylidene)methyl) furan-2-yl)benzoic acid (3f)
Prepared from methyl 4-(5-formylfuran-2-yl)benzoate and com
pound 11 andobtainedasayellowsolid:yield,91%, mp:289e290 ꢀC; IR
(KBr), v(cmꢁ1): 3382, 2980, 1693, 1601, 1358, 1282, 1153; 1H NMR
(DMSO-d6)
d
¼ 7.69(s, 1H), 7.91e7.90(d, J ¼ 6.0 Hz, 1H), 7.72e7.71(d,
J ¼ 6.0 Hz, 1H), 7.47e6.97(m, 12H), 2.25(s, 3H), 2.07(s, 3H); 13C NMR
(DMSO-d6) d 166.6, 165.0, 154.9, 150.7, 150.0, 146.8, 136.0, 134.2, 132.4,
130.6, 130.2, 129.8, 129.2, 126.9, 126.5, 124.8, 123.6, 119.8, 119.8, 116.5,
111.4, 17.4, 17.1; ESI-MS m/z 493.3([M ꢁ H]þ); HRMS calcd for
C29H23N2O4S [M þ H]þ 495.1379, found 495.1375.
5.7. Antibiofilm and antibacterial activity assay
[20] J. Huebner, D.A. Goldmann, Annu. Rev. Med. 50 (1999) 223e236.
[21] Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow
Aerobically; Approved Standard M7-A7, seventh ed. (2006) PA, USA.
The S. epidermidis ATCC35984 was selected as the biofilm-
forming bacteria employed in the study because S. epidermidis is