The antinociceptive evaluation of 2,3-substituted-1,3-thiazolidin-4-ones through thermal stimulation in mice

Article abstract of DOI:10.1007/s00044-017-2052-1

The present study assessed the 2,3-substituted-1,3-thiazolidin-4-ones antinociceptive potential looking at the acute nociception model induced by thermal stimulation in mice. This was done to contribute to the development of new analgesic drugs, in additi

Full text of DOI:10.1007/s00044-017-2052-1

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2018) 27:186–193
DOI 10.1007/s00044-017-2052-1
ORIGINAL RESEARCH
The antinociceptive evaluation of 2,3-substituted-1,3-thiazolidin-4-
ones through thermal stimulation in mice
Arthur H. S. Neves1,2 Daniel S. da Silva1,3 Geonir M. Siqueira1,3
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●
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Giovana D. Gamaro1 Wilson Cunico
Adriana L. da Silva1,2
1,3
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Received: 21 April 2017 / Accepted: 28 August 2017 / Published online: 8 September 2017
© Springer Science+Business Media, LLC 2017
Abstract The present study assessed the 2,3-substituted-
1,3-thiazolidin-4-ones antinociceptive potential looking at
the acute nociception model induced by thermal stimulation
in mice. This was done to contribute to the development of
new analgesic drugs, in addition to the fact that 4-
thiazolidinones are an important scaffold associated with
many pharmacological activities. The synthesized com-
pounds were characterized by GC-MS and NMR of 1H and
13C and administered at a dose of 100 mg/kg hydrochloride
salt (ip). Sodium dipyrone (250 and 500 mg/Kg; ip) and
tramadol hydrochloride (25 and 50 mg/Kg; ip) were used as
positive controls. The hot plate test was done at a tem-
perature of 50 0.1 °C and animals assessed at 30, 60, and
90 min after the drugs were administered. Among the
fourteen compounds tested, nine (5Aa, 5Ab, 5Ac, 5Ad,
5Ba, 5Bb, 5Bd, 5Ea, and 5Fa) showed significant increases
in latency time when compared to saline (negative control)
and compared to sodium dipyrone (500 mg/Kg; ip) in a 30-
min assessment. The highest latency times were obtained
for 3-(2-piperidin-1-yl)ethyl)thiazolidin-4-one derivatives
(5Ab, 5Ac, and 5Ad). This highlights three findings about
the chemical structure that improve activity: (i) an ethylenic
link; (ii) a six-membered piperidine; (iii) an aliphatic sub-
stituent at the 2-position of thiazolidinone ring.
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Keywords Antinociception 4-Thiazolidinones Analgesic
Acute pain Hot plate test
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Introduction
Electronic supplementary material The online version of this article
(https://doi.org/10.1007/s00044-017-2052-1) contains supplementary
material, which is available to authorized users.
In 1979, the International Association for the Study of Pain
defined pain as “an unpleasant sensory and emotional
experience associated with actual or potential tissue damage
or described in terms of such damage,” which is still valid
today (Kopf and Patel 2010). For pain treatment, non-
pharmacological, pharmacological, and invasive methods
are used (Costa et al. 2007; Teixeira et al. 2013; Nossaman
et al. 2010). Thus three classes of analgesic drugs may be
described, opioid analgesic, non-steroidal anti-inflammatory
(NSAIDs), and adjuvant analgesics, such as antidepressant,
anticonvulsant, and anxiolytic medications. The combina-
tion of these analgesic drugs is very useful for chronic pain
therapy (Costa et al. 2007; Liu et al. 2010; Shinde et al.
2015).
* Wilson Cunico
wjcunico@yahoo.com.br
* Adriana L. da Silva
adrilourenco@gmail.com
1
Programa de Pós-Graduação em Bioquímica e Bioprospecção,
Centro de Ciências Químicas Farmacêuticas e de Alimentos,
Universidade Federal de Campus Universitário s/n° cx postal 354,
Pelotas, RS 96010-900, Brazil
2
Departamento de Fisiologia e Farmacologia, Instituto de Biologia,
Universidade Federal de Pelotas, Campus Universitário s/n° cx
postal 354, Pelotas, RS 96010-900, Brazil
3
Laboratório de Química Aplicada a Bioativos, Centro Ciências
The most commonly used method for the relief of acute
pain is systemic analgesics (Camu and Vanlersberghe
2002). However, independently of the type of pain, acute or
Químicas, Farmacêuticas e de Alimentos, Universidade Federal de
Pelotas, Campus Universitário s/n° cx postal 354, Pelotas, RS
96010-900, Brazil

Med Chem Res (2018) 27:186–193
187
chronic, the pharmacological therapy based on opioids and
NSAIDS has several side effects, such as abuse, overdose,
tolerance, dependence, sedation, gastrointestinal dysfunc-
tion, among others. The patients sometimes require other
medication for controlling and managing these side effects
(Bjarnason et al. 1993; Nelson and Camilleri 2015), thereby
decreasing the adherence to pharmacological treatment
(Kurita and Pimenta 2003). Thus, researchers shall develop
new drugs for pain treatment rather safer and more effec-
tive, with side effect reduction that limits medication use,
with greater therapeutic success for the user (Verçoza et al.
2009).
Avance 500 spectrometer (1H at 500 MHz and 13C at 125
MHz) or on a Bruker AC-200F spectrometer (1H at 200
MHz and 13C at 50 MHz) in CDCl3 or D2O containing
tetramethylsilane as an internal standard. The mass spectra
were obtained on a Shimadzu GC-MS-QP2010SE with a
split-splitless injector and equipped with a RDX-SMS
capillary column (30 m × 0.25 mm × 0.25 μm); helium was
used as the carrier gas (56 kPa).
General procedure for the synthesis of thiazolidinones
4Aa–f, 4Ba–d, 4Ca–Fa
In this sense, the 1,3-thiazolidin-4-ones ring has become
very important within the scientific community due to its
multiple biological actions (Jain et al. 2012; Tripathi et al.
2014). Accordingly, researchers have established actions
opposing germs and other infectious agents (virus, bacteria,
fungi and parasites), against chronic diseases such as dia-
betes, hyperlipidemia and convulsions, as well as the pre-
sence of an anti-inflammatory and analgesic effect, among
others (Tripathi et al. 2014; Cunico et al. 2008). Moreover,
1,3-thiazolidin-4-one derivatives have a low cost and large
synthesis versatility, and also large structural variety
depending on the adopted synthesis method. There are
usually substituents at the 2, 3, and 5-position of the ring
that promote changes in chemical, physical, and biological
parameters (Jain et al. 2012; Cunico et al. 2008).
The compounds 4Aa, 4Ac, 4Ae, and 4Af were obtained
according Kunzler et al. (2013). The compounds 4Ea and
4Fa were obtained according Gouvea et al. (2012). For
novel compounds, at first, the reaction occurs through a
solution of 1 mmol of amine 1A–F and 1 mmol of aldehyde
or ketone 2a–f in reflux of toluene (30 mL) for 2 h using a
Dean-Stark trap. After, 3 mmol of mercaptoacetic acid 3
was added and the mixture was heated for more 3 h. The
organic layer was washed with saturated NaHCO3 (3 × 30
mL), dried with MgSO4 and concentrated to give the pro-
ducts that were purified by column chromatography using
hexane:ethyl acetate (7:3) as eluent. The atom numbering
for NMR analyzes identification of compounds 4 and 5 are
given in Fig. 1.
Therefore, taking advantage of the experience of our
group in the synthesis of thiazolidinones and seeking to
contribute to the development of new drugs for pain, the
present study aimed at performing the synthesis of 2,3-
substituted-1,3-thiazolidin-4-ones and antinociceptive
activity through the acute nociception model, induced by
thermal stimulation (hot plate test) in mice.
2-Butyl-3-(2-(piperidin-1-yl)ethyl)thiazolidin-4-one
1
(4Ab) It was obtained as an oil; H NMR δ (CDCl3, 400
MHz, ppm, J H–H = Hz); 4.76 (td, 1H, 3J = 8.5, 4J = 2.3, H-
2); 3.74 (ddd, 1H, 2J = 13.7, 3J = 7.7, 3J = 6.0, H-6a); 3.50
2
4
2
(dd, 1H, J = 15.5, J = 1.5, H-5a); 3.41 (d, 1H, J = 15.5,
2
3
H-5b); 3.04 (dt, 1H, J = 13.9, J = 6.9, H-6b); 2.47 (ddd,
2
3
3
2
1H, J = 13.4, J = 7.3, J = 6.5, H-7a); 2.35 (ddd, 1H, J
= 12.8, 3J = 7.1, 3J = 5.8, H-7b); 2.33–2.39 (m, 4 H);
1.81–1.89 (m, 1H); 1.56–1.63 (m, 1H); 1.46–1.52 (m, 4H);
1.33–1.38 (m, 2H); 1.24–1.32 (m, 4H); 0.82–0.87 (m, 3H).
13C NMR δ (CDCl3, 100 MHz, ppm); 171.0 (C-4); 62.0 (C-
2); 56.1 (C-7); 54.6 (2 C); 39.8 (C-6); 35.2; 32.0 (C-5);
26.3; 25.8 (2C); 24.1; 22.3; 14.0. MS (70 eV): m/z (%) =
270 (M+, 1); 111 (3); 102 (1); 98 (100); 84 (2).
Material and methods
Chemical part
All common reagents and solvents were used and obtained
from commercial suppliers without further purification.
Reactions progress were monitored by a thin-layer chro-
matography (hexane:ethyl acetate 3:1) and/or by a Shi-
madzu Gas Chromatograph GC-2010, HP-1 column (cross
linked methyl siloxane, 30 m × 0.32 mm × 0.25 μm): Col-
umn head pressure, 14 psi, program: T0 = 60 °C; t0 = 2.0
min; rate 10.0 °C min–1; Tf = 280 °C; tf = 13.0 min; Inj. =
250 °C; Det. = 280 °C. The melting points were determined
using open capillaries on a Fisatom model 430 apparatus
and are uncorrected. 1H and 13C nuclear magnetic resonance
(NMR) spectra were recorded on a Bruker DRX 400 spec-
trometer (1H at 400 MHz and 13C at 100 MHz), on a Bruker
2-(4-Fluorophenyl)-3-(2-(pyrrolidin-1-yl)ethyl)thiazolidin-
4-one (4Ba) It was obtained as a solid m.p.: 56–58 °C; 1H
O
O
4
4
5
5
7
7
N
N
N
S
S
N
8
6
6
2
2
A,B
C,D
Fig. 1 The atom numbering for compounds 4 and 5

188
Med Chem Res (2018) 27:186–193
NMR δ (CDCl3, 200 MHz, ppm, J H–H = Hz); 7.26–7.34
(m, 2H, Ar); 7.02–7.12 (m, 2H, Ar); 5.86 (s, 1H, H-2);
3.75–3.88 (m, 2H, H-5a, H-6a); 3.71 (d, 1H, J = 15.5, H-
32.9 (C-5); 22.7 (C-7); 9.3 (2C). MS (70 eV): m/z (%) =
310 (M+, 0.5); 281 (2); 182 (1); 109 (10); 86 (100); 72 (10).
2
5b); 2.65–2.88 (m, 2H, H-6b, H-7a); 2.46–2.60 (m, 5H, H-
7b, H-8); 1.74–1.80 (m, 4H). 13C NMR δ (CDCl3, 50 MHz,
ppm, J C–F = Hz); 171.3 (C-4); 162.9 (d, 1J = 247.7,
Ar),135.3 (d, 4J = 3.2, Ar), 129.1 (d, 2C, 3J = 8.5, Ar),
2-(4-Fluorophenyl)-3-(3-(piperidin-1-yl)propyl)thiazolidin-
4-one (4Da) It was obtained as an oil; 1H NMR δ (CDCl3,
3
4
500 MHz, ppm, J H–H = Hz); 7.23 (dd, 2H, J = 8.6, J =
5.1, Ar); 7.00 (t, 2H, 3J = 8.5, Ar); 5.64 (d, 1H, 4J = 1.7, H-
2); 3.72 (dd, 1H, 2J = 15.6, 4J = 1.8, H-5a); 3.62 (d, 1 H, 2J
= 15.6, H-5b); 3.57 (ddd, 1H, 2J = 14.1, 3J = 8.0, 3J = 6.6,
2
116.0 (d, 2C, J = 21.8, Ar); 63.2 (C-2); 54.0 (2C, C-8);
52.9 (C-7); 41.2 (C-6); 32.9 (C-5); 23.4 (2C). MS (70 eV):
m/z (%) = 294 (M+, 2); 292 (M − 4, 0.5); 224 (0.5); 153
(1); 139 (2); 97 (6); 84 (100); 69 (3).
2
3
3
H-6a); 2.63 (ddd, 1H, J = 13.9, J = 8.0, J = 5.9, H-6b);
2.18–2.23 (m, 5H, H-8a); 2.10–2.16 (m, 1H, H-8b);
1.60–1.66 (m, 1H, H-7a); 1.52–1.57 (m, 1H, H-7b);
1.45–1.49 (m, 4H); 1.34 (sl, 2H). 13C NMR δ (CDCl3, 125
2-Butyl-3-(2-(pyrrolidin-1-yl)ethyl)thiazolidin-4-one
1
1
MHz, ppm, JC–F = Hz); 171.0 (C-4); 162.9 (d, J = 248.0,
(4Bb) It was obtained as an oil; H NMR δ (CDCl3, 400
4
3
MHz, ppm, J H–H = Hz); 4.70 (td, 1H, 3J = 8.5, 4J = 2.3, H-
Ar); 135.3 (d, J = 6.4, Ar); 128.9 (d, 2C, J = 8.2, Ar);
2
2); 3.77 (ddd, 1H, 2J = 14.2, 3J = 8.2, 3J = 6.0, H-6a); 3.51
115.9 (d, 2C, J = 21.8, Ar); 62.9 (C-2); 56.0 (C-8); 54.3
2
4
2
(2C); 41.2 (C-6); 32.8 (C-5); 25.7 (2C); 24.2; 23.9 (C-7).
MS (70 eV): m/z (%) = 322 (M+, 2); 238 (1); 182 (1.5); 127
(4); 112 (4.5); 98 (100); 84 (9).
(dd, 1H, J = 15.5, J = 1.5, H-5a); 3.42 (d, 1H, J = 15.5,
2
3
3
H-5b); 3.10 (ddd, 1H, J = 13.9, J = 7.9, J = 6.0, H-6b);
2.69 (ddd, 1H, 2J = 12.0, 3J = 8.4, 3J = 5.9, H-7a);
2.50–2.56 (m, 5H); 1.82–1.90 (m, 1H); 1.72 (br, 4H);
1.55–1.65 (m, 1H); 1.25–1.31 (m, 4H); 0.84–0.87 (m, 3H).
13C NMR δ (CDCl3, 100 MHz, ppm); 171.1 (C-4); 61.9 (C-
2); 54.1 (2C); 53.0 (C-7); 41.4 (C-6); 35.3; 32.0 (C-5); 26.2;
23.4 (2C); 22.3; 14.0; MS (70 eV): m/z (%) = 256 (M+, 1);
252 (M − 4, 2); 102 (1); 97 (9); 84 (100); 69 (3).
General procedure for the synthesis of hydrochloride salt
5Aa–f, 5Ba–d, and 5Ca–Ga
In a solution of thiazolidinone 4 in dichloromethane (20 ml)
was flowing the hydrochloric acid (HCl (g)) (generate for the
reaction of H2SO4 concentrated with NaCl) in an open
vessel at room temperature for 30 min. The hydrochloride
salt was filtered off under vacuum. When necessary, the salt
was extracted with distilled water from the organic layer.
2-Phenyl-3-(2-(pyrrolidin-1-yl)ethyl)thiazolidin-4-one
1
(4Bc) It was obtained as an oil; H NMR δ (CDCl3, 400
MHz, ppm, J H–H = Hz); 7.27–7.33 (m, 3H, Ar); 7.20–7.24
(m, 2H, Ar); 5.78 (d, 1H, 4J = 1.6, H-2); 3.74 (dt, 1H, 2J =
3
2
4
13.7, J = 6.7, H-6a); 3.72 (dd, 1H, J = 15.5, J = 1.9, H-
4-(2-(Piperidin-1-yl)ethyl)-1-thia-4-azaspiro[4.5]decan-3-
one chlorohydrate (5Ad) It was obtained as a brown solid;
1H NMR δ (D2O, 400 MHz, ppm, J H–H = Hz); 3.59 (t, 2H,
3J = 6.8, H-6); 3.48–3.52 (m, 2H); 3.48 (s, 2H, H-5); 3.12
5a); 3.64 (d, 1H, 2J = 15.4, H-5b); 2.72–2.79 (m, 2H, H-6b,
2
3
H-7a); 2.61 (dt, 1H, J = 13.5, J = 6.8, H-7b); 2.37–2.40
(m, 4H); 1.65–1.69 (m, 4H). 13C NMR δ (CDCl3, 100 MHz,
ppm); 171.3 (C-4); 139.6 (Ar); 129.1 (Ar); 129.0 (2C, Ar);
127.0 (2C, Ar); 64.0 (C-2); 54.1 (2C); 53.1 (C-7); 41.6 (C-
6); 32.9 (C-5); 23.4 (2C); MS (70 eV): m/z (%) = 276 (M+,
1); 272 (M-4, 2); 178 (0.5); 135 (2); 121 (3); 97 (10); 84
(100); 70 (3).
3
2
4
(t, 2H, J = 6.8, H-7); 2.83 (dt, 2H, J = 12.4, J = 2.3);
1.74–1.83 (m, 4H); 1.63–1.70 (m, 5H); 1.28–1.60 (m, 6H);
0.93–1.04 (m, 1H); 13C NMR δ (D2O, 100 MHz, ppm);
175.1 (C-4); 75.2 (C-2); 55.8 (C-7); 53.9 (2C); 37.1 (2C);
36.4 (C-6); 30.6 (C-5); 23.7; 23.0 (2C); 22.9 (2C); 21.0. MS
(70 eV) (free base 4Ad): m/z (%) = 171 (M+-111, 0.5); 128
(2); 111 (6); 98 (100); 84 (2).
3-(3-(Diethylamino)propyl)-2-(4-fluorophenyl)thiazolidin-
4-one (4Ca) It was obtained as an oil; 1H NMR δ (CDCl3),
3
4
600 MHz, ppm, JH–H = Hz); 7.35 (dd, 2H, J = 8.6, J =
4-(2-(Pyrrolidin-1-yl)ethyl)-1-thia-4-azaspiro[4.5]decan-3-
one chlorhydrate (5Bd) It was obtained as a dark brown
solid; 1H NMR δ (D2O, 400 MHz, ppm, J H–H = Hz);
3.60–3.66 (m, 2H); 3.58 (t, 2H, 3J = 6.8, H-6); 3.49 (s, 2H,
H-5); 3.25 (t, 2H, 3J = 6.7, H-7); 2.96–3.03 (m, 2H);
1.94–2.06 (m, 2H); 1.83–1.91 (m, 2H); 1.75–1.82 (m, 2H);
3
5.2, Ar); 7.08 (t, 2H, J = 8.5, Ar); 5.75 (s, 1H, H-2); 3.79
2
4
2
(dd, 1H, J = 15.6, J = 1.5, H-5a); 3.72 (d, 1H, J = 15.6,
H-5b); 3.62 (dt, 1H, 2J = 14.2, 3J = 7.6, H-6a); 2.78 (q, 4H,
3J = 7.2); 2.74–2.76 (m, 1H, H-6b); 2.65–2.70 (m, 1H, H-
8a); 2.59–2.63 (m, 1H, H-8b); 1.75–1.82 (m, 2H, H-7); 1.12
(t, 6H, 3J = 7.2). 13C NMR δ (CDCl3, 150 MHz, ppm, JC–F
2
1.64–1.71 (m, 4H); 1.49 (d, 1H, J = 13.0); 1.32–1.44 (m,
1
4
= Hz); 171.5 (C-4); 163.0 (d, J = 248.8, Ar); 134.9 (d, J
2H); 0.93–1.05 (m, 1H). 13C NMR δ (D2O, 100 MHz,
ppm); 175.0 (C-4); 75.2 (C-2); 54.8 (2C, C-8); 54.0 (C-7);
37.8 (C-6); 37.0 (2C); 30.6 (C-5); 23.7; 23.1 (2C); 22.6
3
2
= 3.1, Ar); 129.2 (d, 2C, J = 8.4, Ar); 116.1 (d, 2C, J =
21.9, Ar); 62.7 (C-2); 49.1 (C-8); 45.6 (2C); 40.6 (C-6);

Med Chem Res (2018) 27:186–193
189
(2C). MS (70 eV) (free base 4Bd): m/z (%) = 264 (M+-4,
3); 171 (1); 128 (1); 97 (15); 84 (100); 69 (3).
responses, the animals were disposed from the next stage
(test).
In the day of the test, the mice were placed separately on
the apparatus, and the latency time (in seconds) for the
nociceptive response (jumping or hind paw licking) was
measured with a manual chronometer in times of 30, 60,
and 90 min after the saline injection (NaCl 0.9%, 10 ml/Kg,
ip), 2,3-substituted-1,3-thiazolidin-4-ones derivatives (100
mg/Kg, ip), dipyrone (250 and 500 mg/Kg, ip) or tramadol
(25 and 250 mg/Kg, ip); it was settled the upper cut-off time
of 50 s, to avoid possible tissue damage in animals. The
temperature established in the experiment was set to 50
0.1 °C.
Biological part
Animals
Experiments were performed using 60–90 day-old adult
male (Swiss) mice. Animals were maintained in controlled
environmental conditions (22 1 °C, 12/12 h light/dark
cycle, relative humidity (45–55%) and free access to food
and water (Luszczki 2010). All experiments were based on
precepts and ethical considerations, to investigate experi-
mental pain in animals (Zimmermann 1983). The project
was approved by the University Ethics Committee, regis-
tered under the number (CEEA 2231).
Finally, the animals were submitted to the hot plate test
for three different moments after treatment administration,
in 30, 60, and 90 min, and the latency time data were
registered for further analysis (Vigorita et al. 2001).
Statistical analysis
Standard drugs and negative control
Data were analyzed by using variance analysis (ANOVA)
followed by Duncan's test in the SPSS 11.0.1 software.
Furthermore, all data were expressed as mean standard
error and the level of significance was set as P < 0.05.
Sodium dipyrone (Novalgina®, Aventis Pharma Ltd) was
diluted in water at two concentration 250 and 500 mg/Kg.
Tramadol hydrochloride (Tramal®, Pfizer Ltd) was diluted
in water at two concentration 25 and 50 mg/Kg. Saline
solution (sodium chloride 0.9%, Equiplex Ltd) was used as
a negative control. Vehicle and drugs were administered
intraperitoneally in volume of 0.1 ml/10 g body weight.
Results and discussion
Chemistry
Hot plate test
Fourteen 1,3-thiazolidin-4-ones, eight of them unpublished
in the literature, were synthesized for biological study. The
new ones and the compounds 4Aa (Kunzler et al. 2013),
4Ac (Sun and Kyle 2003), 4Ae (Kunzler et al. 2013), 4Af
(Kunzler et al. 2013), 4Ea (Gouvêa et al. 2012), and 4Fa
(Gouvêa et al. 2012) were synthesized from reactions
between amines 1A–F with aldehydes or ketone 2a–f in
toluene reflux. Scheme 1 shows the conditions to the
synthesis of desired compounds.
According to many studies in literature, the anti-
nociceptive activity of some 1,3-thiazolidin-4-ones might
be associated with their capacity to inhibit cyclooxygenase
(COX) enzymes including COX-2 selective inhibition,
(Geronikaki et al. 2008; Taranalli et al. 2008; Unsal-Tan
et al. 2012; Vigorita et al. 2003; Zarghi et al. 2007). Enzyme
inhibition shows anti-inflammatory, analgesic and anti-
pyretic activities due to the reduction of prostaglandin
synthesis (Davis and Brogden 1994). In this way, a preview
study showed the analgesic and anti-inflammatory activities
The hot plate test is a standard model test used to determine
antinociceptive efficacy of drugs with central activity
through acute thermal stimulation (Siegfried et al. 1987).
The apparatus used (Hot Plate, model EFF361, Insigth®)
consisted of an aluminum plate that is evenly heated, and it
is surrounded by a transparent acrylic rectangular cage,
which keeps animals confined, where access is done only by
a higher opening that allows researchers to remove and
include animals.
Animals were randomized into groups of eight subjects
and were habituated in a room for at least 30 min before the
experiments (Luszczki 2010). Twenty-four hour before
each experiment, animals were weighed and accustomed to
procedure, to avoid the occurrence of new-induced
analgesia which could provide false results (Siegfried
et al. 1987). In habituation, the mice were exposed to the
same conditions that would be subject on the day of the
experiment, except to the derivatives treatment, such as the
researcher manipulation, the injection (saline) and the mice
insertion in the equipment. Lastly, the animals were
observed, for a minute, for their pain responses and then
they were removed from the equipment. If positive for pain
of
2-(aryl)-3-(4-sulfonamidebenzene)-1,3-thiazolidin-4-
ones. The compound with 4-fluorophenyl as an aryl group
showed good results compared to NSAID nimesulide
(Prasit et al. 1999). In this sense, this study began

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Med Chem Res (2018) 27:186–193
Scheme 1 Synthesis of
thiazolidinones 4 and their
hydrochloride salts 5
researching 2-(4-fluorophenyl)-thiazolidinones (4Aa–Ga)
with different amine cores (A–F) linked at 3-position at the
thiazolidinone ring (Scheme 1).
Thiazolidinones 4Ab–f and 4Bb–d were synthesized
according to biological results. For this, the amines A (2-
(piperidin-1-yl)ethan-1-amine) and B (2-(pyrrolidin-1-yl)
The 1,3-thiazolidin-4-ones were identified by classical
H-2 and H-5 signals at H NMR. Due to the presence of a
1
stereogenic center at 2-position, the H-5 hydrogens are
diastereotopic and present two signals: one at ~3.5 ppm for
2
4
H-5a (doublet of doublet with J = 15.5 Hz and J = 1.5
2
Hz); and other at ~3.4 ppm for H-5b (doublet with J =
ethan-1-amine)
were
maintained
and
the
4-
15.5 Hz). The H-2 was assigned as a simplet or doublet at
~5.7 ppm, with the exception of thiazolidinones from
valeraldehyde (4Ab and 4Bb) those H-2 appear as triplet of
doublet at ~4.7 ppm. For thiazolidinone salt from cyclo-
hexanone (5Ad and 5Bd), the both H-5 appear as simplet at
~3.4 ppm since these compounds have no stereogenic center
at 2-position.
fluorobenzaldehyde (a) was modified in the synthetic pro-
cess, obtaining compounds based on other carbonyl group
(aldehydes and cyclohexanone, b–f).
In sequence, all thiazolidinones underwent a reaction to
form a hydrochloride salt (5) in the basic nitrogen that all
amines have in their structures. Due to salt formation, no
surfactant agent was needed to apply it in an animal model.
Efficient methodologies were studied at aiming to obtain
all all the proposed products in moderate to good yields
(Table 1). The structure of known thiazolidinones (4Aa,
4Ac, 4Ae, 4Af, 4Ea, and 4Fa) was confirmed by mass
spectrometry. The new compounds were fully identified and
characterized by gas chromatography-mass spectrometry
and 1H and 13C NMR. The NMR of compounds 4Ab, 4Ba,
4Bb, 4Bc, 4Ca, and 4Da was performed for the free base
and compounds 5Ad and 5Bd for the hydrochloride salt.
At 13C NMR, the mainly signals for thiazolidinones 4
are: C-4 at ~170 ppm; C-2 at ~62 ppm; and C-5 at ~32 ppm.
Compounds 5 have a deshield C-2 at ~75 ppm and a
shielded C-5 at ~30 ppm.
Antinociceptive study
The hydrochloride salts 5 were tested in an animal model. In
the hot plate test, pain response is described as latency time
(in seconds), thus the high latency times indicate good

Med Chem Res (2018) 27:186–193
191
Table 1 Chemical parameters
of 2,3-substituted-1,3-
thiazolidin-4-ones
Yield (%)
MF
MW (g/mol)
Log P*
Hydrochloride salt (5)
MW (g/mol)
Dose (µmol/kg)**
4Aa
4Ba
4Ca
4Da
4Ea
4Fa
4Ab
4Ac
4Ad
4Ae
4Af
4Bb
4Bc
4Bd
74
89
22
52
44
59
64
68
48
33
71
62
66
15
C16H21FN2OS
308.41
294.39
310.43
322.44
288.34
274.31
270.43
290.42
282.44
335.42
320.45
256.41
276.40
268.42
2.72
2.30
2.77
2.83
2.81
3.03
2.31
2.56
2.33
2.52
2.44
1.90
2.14
1.91
344.87
330.85
346.89
358.90
324.80
310.77
306.89
326.88
318.90
371.88
356.91
292.87
312.86
304.88
289.9
302.2
288.2
278.6
307.8
321.7
325.8
305.9
313.5
268.9
280.2
341.4
319.6
328.0
C
C
C
C
C
15H19FN2OS
16H23FN2OS
17H23FN2OS
15H13FN2OS
14H11FN2OS
C14H26N2OS
C
C
C
C
C
C
C
16H22N2OS
15H26N2OS
16H21FN3O3S
17H24FN2O2S
13H24N2OS
15H20N2OS
14H24N2OS
FM molecular formula, MW molecular weight
*Log P calculated by software Chendraw® Ultra, version 8.0.3
**Dose in µmol/Kg corresponding to 100 mg/Kg of hydrochloride salt (5)
50
45
40
#
50
#
#
40
*
*
#
35
*
*
30
30
20
10
0
*
*
*
*
25
*
20
15
10
5
0
NaCl 5Ab 5Ac 5Ad 5Ae 5Af 5Bb 5Bc 5Bd DIP TRA
NaCl
5Aa
5Ba
5Ca
5Da
5Ea
5Fa
DIP
TRA
Fig. 3 Latency time of 3-(piperidin-1-yl)-ethyl-2-substituted-1,3-
thiazolidin-4-ones (5Ab–f) and 3-((pyrrolidin-1-yl)-ethyl)-2-sub-
stituted-1,3-thiazolidin-4-ones (5Bb–d) in hot plate test evaluation in
30 min (100 mg/Kg, ip). DIP Sodium dipyrone (500 mg/Kg); TRA
Tramadol hydrochloride (50 mg/Kg). *P < 0.05 X saline; # P < 0.05 X
negative control and compounds. ANOVA/DUNCAN
Fig. 2 Latency time of 3-amino-2-(4-fluorophenyl)-1,3-thiazolidin-4-
ones (5Aa–Ga) in hot plate test evaluation in 30 min (100 mg/Kg, ip).
DIP Sodium dipyrone (250 mg/Kg); TRA Tramadol hydrochloride (25
mg/Kg). *P < 0.05 X saline; # P < 0.05 X negative control and
compounds. ANOVA/DUNCAN
inhibition of nociception. The latency times of thiazolidin-
4-ones in the hot plate test are shown in Figs. 2, 3. Gen-
erally, the findings agree with others studies in the field,
evidencing, once again, that 1,3-thiazolidin-4-ones are an
important scaffold associated with many pharmacological
activities (Tripathi et al. 2014). Under these circumstances,
from fourteen compounds tested, nine showed significant
increases in latency time when compared to saline (negative
control).
Figure 2 exhibits the effect of compounds 5Aa–Ga on
latency time 30 min after injection. Four (5Aa, 5Ba, 5Ea,
and 5Fa) revealed an increase in latency to nociceptive
response when compared to saline (P < 0.05) by ANOVA/
DUNCAN, demonstrating that these compounds are
capable of promoting antinociceptive activity. However,
none was better than dipyrone or tramadol, two well-known
analgesic drugs. These standard drugs were selected as
positive controls, due to their strong analgesic effect, low
cost and greater current therapeutic use (Teixeira et al.
2013; Silva and Moraes 2010, Edwards et al. 2010; Tulunay
et al. 2004). Although morphine is the standard drug to
evaluate analgesic medication, tramadol hydrochloride was
chosen due to its greater use to relieve acute or chronical
pain of moderate or severe intensity (Silva and Moraes
2010; Teixeira et al. 2013; Giraldes et al. 2016). Never-
theless, to maintain the same analgesic effect, morphine
(mg) was converted to tramadol (mg), using tables of
equianalgesic opioid doses in parenteral administration

192
Med Chem Res (2018) 27:186–193
(1 mg morphine = 10 mg of tramadol) (Kopf and Patel
2010).
nociceptive response compared to dipyrone (33.0 4.6 s).
When evaluating these compounds in 60 and 90 min, none
showed analgesic effect (see Supplementary material).
Moreover, literature findings demonstrated that 1,3-thiazo-
lidin-4-one derivatives have a similar anti-inflammatory and
analgesic efficacy to NSAIDs (Geronikaki et al. 2008;
Vigorita et al. 2001).
The difference in increases of latency time among 5Aa,
5Ba, 5Ea, 5Fa, and dipyrone possibly occurred due to the
dose used. Sodium dipyrone (M.W. 333.3 g/mol; 250 mg/
kg, 750 µmol/Kg; ip) had a molar mass similar to 3-
(amino)-2-(4-fluorophenyl)-1,3-thiazolidin-4-ones 5Aa–Ga
(M.W. 310.81–358.94 g/mol; 100 mg/kg, 278.6–323.7
µmol/Kg; ip). The 100 mg/kg dose was based on previous
works (Taranalli et al. 2008; Vigorita et al. 2001).
In the 60 and 90 min evaluation (see supplementary
information), there were decreases in latency times, which
were expected except for compound 5Ea that still main-
tained higher levels of antinociceptive activity than saline
over these times. This suggests that the compounds tested
have a short effect and fast metabolization, but further
studies are necessary to confirm this hypothesis. Yet a
previous study investigated the anti-inflammatory and
analgesic activity of 3,3′-(1,2-ethanediyl)-bis[2-aryl-4-thia-
zolidinone], obtaining good results and latency times in a
hot plate test as phenylbutazone and indomethacin did,
within up to 180 min after treatment. These compounds
have two 1,3-thiazolidin-4-one nuclei connected by an ethyl
bridge (position 3) and two substituted phenyl at 2-position
(Vigorita et al. 2001).
Based on the results shown in Fig. 2, a 30-min assess-
ment was chosen in the sequence of the study in order to
plan a new structural modification in the two compounds
with the highest latency times: 2-(4-fluorophenyl)-3-
(piperidin-1-ylethyl)-1,3-thiazolidin-4-one (5Aa; 24.3
5.6 s); and 2-(4-fluorophenyl)-3-(pyrrolidin-1-ylethyl)-1,3-
thiazolidin-4-one (5Ba; 20.3 2.6 s). Similarly to Vigorita
et al. (2001), our work showed good results for compounds
with ethyl as a bridge (A and B). In this way, this distance
between the atoms of nitrogen may seem important for the
antinociceptive activity since the increase of chain showed
no statistical activity (propyl link, 5Ca and 5Da). Com-
pounds with shorter distances, methyl (5Ea) or none (5Fa),
also showed good results in 30 min (Fig. 2). Therefore,
comparing compounds with different links between nitro-
gens, the order of activity was: ethyl > methyl or none »
propyl.
It is highlighted that all compounds with six-membered
piperidine (5A) showed better results than compounds with
five-membered pyrrolidine (5B) comparing the same 2-
position substituent (5Aa–d compared to 5Ba–d, Figs. 2,
3). These results suggested that, in addition to an ethylenic
link, a six-member ring is also important to activity.
Another important finding is the influence of substituents
R1 and R2 on antinociceptive activity. Comparing the
compounds with aromatic substituents, phenyl (c) has better
antinociceptive activity than aryl, no matter what the nature
of the aryl substituent, since both electron-withdrawing
fluoro (a) or nitro (e) and electron-releasing methoxy (f)
decrease the activity. The best results were found for
compounds bearing an aliphatic substituent, from valer-
aldehyde (5Ab) and from ciclohexanone (5Ad). These
results suggested that an aliphatic group is also important
for this activity. Comparing compounds with different
groups at 2-position of thiazolidinone ring, the order of
activity was: aliphatic > phenyl > aryl.
Conclusion
In conclusion, this work described the antinociceptive effect
of 1,3-thiazolidin-4-ones through thermal stimulation in
mice. The 1,3-thiazolidin-4-ones were synthesized from
one-pot or multicomponent procedures using different
amines 1A–F and aldehydes or ketone 2a–f. Fourteen
compounds were obtained in moderate to good yields,
identified and characterized. Nine thiazolidinones showed a
significant increase in latency time in the hot plate test.
Three of them (5Ab, 5Ac, and 5Ad) showed the best
antinociceptive activity after 30 min at 100 mg/Kg. It can
highlight three findings about chemical structure: (i) the
distance between nitrogens is important to activity and the
best result was found with an ethylenic link; (ii) the pre-
sence of six-membered heterocycle piperidine is more
efficient; (iii) an aliphatic substituent at 2-position of thia-
zolidinone improves the activity. In addition there are
ongoing anti-inflammatory and anti-pyretic studies as well
as the evaluation of the antinociceptive mechanism.
Further, analogs from A (5Ab–f) and B (5Bb–d) were
studied. Compounds 5Ab, 5Ac, 5Ad, 5Bb, and 5Bd (five of
eight) revealed increased latency times when compared to
saline in the 30-min evaluation after treatment (Fig. 3). The
highest latency times were obtained by 2-butyl-3-(2-piper-
idin-1-yl)ethyl)thiazolidin-4-one (5Ab; 32.4 3.5 s), 2-
phenyl-3-(2-piperidin-1-yl)ethyl)thiazolidin-4-one
(5Ac;
27.8 4.3 s) and 4-(2-piperidin-1-yl)ethyl)-1-thia-4-azas-
piro[4.5]decan-3-one (5Ad; 31.6 4.3 s). These com-
pounds have better antinociceptive activity than 5Aa (24.3
Acknowledgements The authors thank UFPel and FAPERGS (proc.
11/2068-7) for financial support. Fellowships granted to A.H.S.N. by
CAPES, D.S.S by CAPES/FAPERGS and W.C. by CNPq (proc.
308791/2015-0).
5.6 s, 4-fluorophenyl substituent) and showed
a

Med Chem Res (2018) 27:186–193
193
Compliance with ethical standards
Prasit P, Wang Z, Brideau C, Chan C-C, Charlson S, Cromlish W,
Ethier D, Evans JF, Ford-Hutchinson AW, Gauthier JY, Gordon
R, Guay J, Gresser M, Kargman S, Kennedy B, Leblanc Y, Leger
S, Mancini J, O’Neal GP, Quellet M, Percival MD, Perrier H,
Riendeau D, Rodger I, Tagari P, Therien M, Vikers P, Wong E,
Xu L-J, Young RN, Zamboni R, Boyce S, Rupniak N, Forrest M,
Visco D, Patrcik D (1999) The discovery of rofecoxib, [MK 966,
Vioxx, 4-(4’-methylsulfonylphenyl)-3-phenyl-2(5H)-furanone],
an orally active cyclooxygenase-2-inhibitor. Bioorg Med Chem
Lett 9:1773–1778
Conflict of interest The authors declare that they have no competing
interests.
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