Design, synthesis and evaluation of novel thiazolidinedione derivatives as anti-hyperglycemic and anti-hyperlipidemic agents

Article abstract of DOI:10.1007/s00044-016-1675-y

A novel series of thiazolidine-2,4-diones was designed, synthesized and investigated for anti-diabetic activity. The (2,4-dioxo-1,3-thiazolidin-5-yl)methylphenylbenzamide derivatives contain an amide linkage between the central aryl ring and the hydrophobic tail. Structures of the compounds were confirmed by spectroscopic techniques fourier transform infrared spectroscopy, ¹H-nuclear magnetic resonance and elemental (C, H, N) analysis. The synthesized compounds were evaluated for their in-vivo pharmacological activity (blood glucose and triglyceride lowering activity), where compounds thiazolidinediones-C and thiazolidinediones-E showed significant effects, comparable to the standard pioglitazone. Computational studies positively substantiated the nature and interaction of the designed compounds with peroxisome proliferator-activated receptor gamma.

Full text of DOI:10.1007/s00044-016-1675-y

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-016-1675-y
ORIGINAL RESEARCH
Design, synthesis and evaluation of novel thiazolidinedione derivatives
as anti-hyperglycemic and anti-hyperlipidemic agents
1 ●
1 ●
2 ●
1 ●
Sushant K. Shrivastava Ankit Batham Saurabh K Sinha Tanmaya K. Parida
1 ●
1
Debapriya Garabadu Priyanka K. Choubey
Received: 23 January 2016 / Accepted: 4 July 2016
©
Springer Science+Business Media New York 2016
Abstract A novel series of thiazolidine-2,4-diones was
designed, synthesized and investigated for anti-diabetic
activity. The (2,4-dioxo-1,3-thiazolidin-5-yl)methylphe-
nylbenzamide derivatives contain an amide linkage between
the central aryl ring and the hydrophobic tail. Structures of
the compounds were confirmed by spectroscopic techniques
Introduction
Diabetes mellitus (DM) is one of the most common meta-
bolic disorders in the world (Eckel et al., 2005).
According to WHO guidelines, about 347 million people
suffer from DM, and about 80 % of those live in low and
middle income countries. Reports project that this pandemic
will become the seventh leading cause of mortality world-
wide (Wild et al., 2004; Mathers and Loncar, 2006; Kokil
et al., 2015). It has been reported that about 80 % of the
diabetic population have type-2 or non-insulin-dependent
diabetes mellitus (T2DM) (Kopelman and Hitman, 1998;
Mlinar et al., 2007; Gaur et al., 2014) and it is projected that
this proportion will grow by 5.4 % by 2025 (Kim et al.,
2006). T2DM is considered to be a multifaceted and com-
plex etiological disease coupled with several co-morbidities
such as dyslipidemia, atherosclerosis and obesity (Kahn
et al., 1996). Insulin resistance and deficiency are also
phenotypic characteristics for this condition (De Fronzo and
Tripathy, 2009). Several drugs have been developed for the
management of T2DM, however, their use is limited due to
their serious unwanted side effects (Kokil et al., 2015). It is,
therefore, necessary to develop agents that enhance insulin
sensitivity with minimal hypoglycemic episodes in the
pharmacotherapy of T2DM.
1
fourier transform infrared spectroscopy, H-nuclear mag-
netic resonance and elemental (C, H, N) analysis. The
synthesized compounds were evaluated for their in-vivo
pharmacological activity (blood glucose and triglyceride
lowering activity), where compounds thiazolidinediones-C
and thiazolidinediones-E showed significant effects, com-
parable to the standard pioglitazone. Computational studies
positively substantiated the nature and interaction of the
designed compounds with peroxisome proliferator-activated
receptor gamma.
●
●
●
Keywords Thiazolidindione PPAR γ Pioglitazone
Insulin sensitizer
Electronic supplementary material The online version of this article
doi:10.1007/s00044-016-1675-y) contains supplementary material,
which is available to authorized users.
Thiazolidine-2,4-diones are a class of prototypical insulin
sensitizer widely used for the treatment of T2DM. These
drugs control carbohydrate and lipid metabolism through a
class of ligand-activated nuclear peroxisome proliferator-
activated receptors (PPARs) (Dumasia et al., 2005). These
drugs are reported to attenuate insulin resistance without
exhibiting hypoglycemia (Kokil et al., 2015; Skyler, 2004;
Fujita et al., 1983). Drugs in this class, including rosiglita-
zone, pioglitazone, ciglitazone, troglitazone and englitazone,
exhibit significant anti-diabetic activity. However, drugs like
(
*
Sushant K. Shrivastava
skshrivastava.phe@iitbhu.ac.in
1
Pharmaceutical Chemistry Research Laboratory, Department of
Pharmaceutics, Indian Institute of Technology (BHU), Varanasi
2
21 005, India
2
Department of Pharmaceutical Sciences, Mohanlal Sukhadia
University, Udaipur 313001, India

Med Chem Res
Hydrophobic tail
Linker 2
H
O
S
Cl
N
a
O
O
+
S
HO
H
N
2
NH
2
Linker 1
b
H
N
S
O
Thiazolidinedione Moiety
NO2
NH2
NH
O
O
O
O
S
S
c
Central aryl ring
HN
HN
Fig. 1 Different regions of the designed molecule
O
O
Intermediate II
Intermediate I
rosiglitazone exert unwanted side effects such as hepato-
toxicity and genotoxicity (Scheen, 2001; Bedir et al., 2006).
Hence, there is a need to develop novel thiazolidinedione
derivatives that have equal or better therapeutic effect and
minimal toxicity. To this end, several strategies have been
attempted to reduce peripheral toxicity in comparison with
established thiazolidinedione drugs in the market.
In established anti-diabetic thiazolidinediones (TZDs), the
hydrophobic tail is substituted with a range of aryl, heteroaryl
and cycloalkyl rings (Yanagisawa et al., 2000; Hulin et al.,
O
R
d
Cl
R
NH
O
O
HN
S
O
Scheme 1 Synthesis of thiazolidinedione derivatives (a) thiourea,
HCl, H O, 8–10 h reflux at 100–110 °C. (b) piperidine, toluene, ben-
zoic acid, 2-h reflux. (c) CoCl .6H O, dimethylglyoxime, H O, NaOH,
NaBH , Tetrahydrofuran (THF). (d) NaOH
2
1
992), and the central phenyl group is substituted with other
aromatic rings (Bernardon and Clary, 2002; Cantello et al.,
994). A series of imidazopyridine thiazolidine-2,4-dione
2
2
2
4
1
derivatives of rosiglitazone are reported to possess anti-
diabetic activity with reduced cardiotoxicity (Oguchi et al.,
JEOL, AL300, FT-NMR spectrophotometer with operating
frequency, 300 MHz at temperature 25 C using dimethyl
o
2
000). 5-Benzylidene-2,4-thiazolidinedione analogs are
sulfoxide (DMSO)-d6 as solvent containing Tetra-
methylsilane as internal standard.
reported to have anti-hyperglycemic and anti-hyperlipidemic
activity (Munj et al., 2011).
Based on these structural modifications, the present study
aims to design and synthesize specific thiazolidinedione
analogs with an amide linkage instead of an ether linkage
between the lipophilic tail and the central phenyl ring
Chemistry
Thiazolidinedione was synthesized by reacting chloroacetic
acid with thiourea in the presence of HCl and water as
shown in Scheme 1 (Parekh et al., 2013). The resulting
thiazolidine-2,4-dione was treated with 4-nitrobenzaldehyde
in the presence of piperidine, toluene and benzoic acid to
give 5-(4-nitrobenzyliden)thiazolidine-2,4-dione (Inter-
mediate I; Storr et al., 2003). This was then selectively
reduced to give 5-(4-aminobenzyl)thiazolidine-2,4-dione
(
linker 2) as shown in Fig. 1.
Materials and methods
Materials
(Intermediate II; Cantello et al., 1994). Finally Intermediate
All reagents and solvents were of analytical grade and
purchased from Merck and Sigma Aldrich. Streptozotocin,
nicotinamide and pioglitazone were procured from Sigma
Aldrich. All reactions were monitored by thin layer chro-
matography, using iodine as an indicator. Open capillary
melting point apparatus was used to detect melting points
and are uncorrected. fourier transform infrared spectroscopy
II was treated with a range of substituted benzoyl chlorides
to afford the desired compounds (2,4-dioxo-1,3-thiazolidin-
5-yl)methylphenylbenzamides (Furniss et al., 1989).
Synthesis of thiazolidinediones
An 6 mL aqueous solution of 5.6 g of chloroacetic acid was
added to an 6 mL aqueous solution of 4.56 g of thiourea and
stirred for 15 min, resulting in a white precipitate accom-
panied by considerable cooling. The resulting precipitate
was treated with conc. HCl and refluxed for 10 h at
100–110 °C with stirring. The mixture was then cooled and
(
FT-IR) spectra were recorded using a KBr pellet method in
a SHIMADZU Infrared-spectrophotometer, FTIR-8400S.
Elemental analysis was performed with an Exeter Analy-
1
tical Inc., CE-440 elemental analyzer (USA). H nuclear
magnetic resonance (NMR) spectra were measured by

Med Chem Res
the resulting white needles were filtered, washed with water
and recrystallized from ethanol.
benzamide), 135.7 (CH, Ar-C, C-1’), 135.9 (CH, Ar-C,
C-4’), 121.3 (2 × CH, Ar-C, C-2’, C-6’), 127.8 (2 × CH,
Ar-C, C-3’, C-5’), 131.4 (CH, Ar-C, C-1”), 147.5 (CH, Ar-C,
C-4”), 127.5 (2 × CH, Ar-C, C-2”, C-6”), 127.6 (2 × CH,
Synthesis of 5-(4-nitrobenzylidine)thiazolidine-2,4-dione
(
Intemidiate I)
Ar-C, C-3”, C-5”), 56.9 (CH, C-5), 35.6 (CH , CH linker),
2 2
2
8.7 (CH ,CH CH ), 15.9 (CH , CH CH ); Anal. Calcd.
2
2
3
3
2
3
A mixture of 2.5 g of 4-nitrobenzaldehyde and 1.9 g of
thiazolidin-2,4-dione in 10 mL of toluene was treated with
for C19H N O S: C, 64.3; H, 5.1; N, 7.9. Found: C, 63.95;
18 2 3
H, 5.03; N, 8.15.
0
.3 g of benzoic acid and 0.28 mL of piperidine. The reac-
N-(4-((2,4-dioxothiazolidin-5-yl)methyl)phenyl)-4-methox-
ybenzamide [TZDs-M] This compound was synthesized
from Intemidiate II and 4-methoxybenzoyl chloride fol-
lowing the same procedure TZDS-E (Yield 89 %) mp
tion mixture was refluxed for 2 h. The Dean-Stark water
separator was used for continuous removal of water. The
mixture was cooled to room temperature and the resulting
precipitate was filtered and washed with dichloromethane
and diethyl ether.
272–275 °C; UV (DMSO)λmax 315; IR (KBr) vmax 3431,
3203, 2924, 1714, 1406, 1151; 1H NMR (DMSO-d6,
500 MHz,): δ = 10.01 (1H, s, NH of thiazolidine ring), 9.05
Synthesis of 5-(4-aminobenzyl) thiazolidine-2,4-dione
(1H, s, NH of benzamide), 6.94–7.84 (8H, Ar-H), 4.22 (1H,
(
Intemidiate II)
t, J = 8.5 MHz, C-H of thiazolidine ring), 3.49(2H, dd,
J = 7 MHz, 12 MHz, CH linker), 3.78 (3H, s,OCH ); 13C
2
3
A solution of 0.3 g of cobalt chloride hexahydrate and 0.6 g
of dimethylglyoxime in 300 mL of water was treated with
3
cooled to 0 °C in an ice bath and treated with 7.9 g of
sodium borohydride, followed by slow addition of a slurry
of 5-(4-aminobenzyl) thiazolidine-2,4-dione in 50 mL of
THF. The resulting mixture was stirred overnight at room
temperature. The pH of the reaction mixture was adjusted to
NMR (DMSO-d6, 125 MHz,): δ = 178.7 (C, C=O, C-2),
1
1
76.8 (C, C=O, C-4), 169.4 (C, CONH of benzamide),
35.09 (CH, Ar-C, C-1’), 134.9 (CH, Ar-C, C-4’), 127.3
mL of 1M NaOH. The resulting brown solution was
(
5
(
5
2 × CH, Ar-C, C-2’, C-6’), 121.4 (2 × CH, Ar-C, C-3’, C-
’), 126.9 (CH, Ar-C, C-1”), 164.5 (CH, Ar-C, C-4”), 128.5
2 × CH, Ar-C, C-2”, C-6”), 119.2 (2 × CH, Ar-C, C-3”, C-
”), 58.9 (CH, C-5), 55.2 (CH , OCH ), 39.6 (CH , CH
2
3
3
2
linker); Anal. Calcd. for C H N O S: C, 60.6; H, 4.5; N,
1
8 16 2 4
9
.0 using acetic acid and the mixture was extracted with
7.8. Found: C, 61.77; H, 4.33; N, 7.66.
THF (3 × 200 mL). The organic extract was dried over
sodium sulfate, concentrated and the residue was recrys-
tallized with ethanol.
4-(chloromethyl)-N-(4-((2,4-dioxothiazolidin-5-yl)methyl)
phenyl)benzamide [TZDs-C] This compound was synthe-
sized from Intemidiate II and 4-chloromethylbenzoyl chlor-
ide following the same procedure TZDS-E (Yield 87 %)
mp 281–285 °C; UV (DMSO)λmax 254; IR (KBr) vmax
Synthesis of different derivatives of [{(2,4-dioxo-1,3-
thiazolidin-5-yl)methyl}phenyl]benzamide
3
5
1
4
454, 3200, 2924, 1714, 1406, 1151; 1H NMR (DMSO-d6,
00 MHz,): δ = 12.58 (1H, s, NH of thiazolidine ring),
0.82 (1H, s, NH of benzamide), 7.10–7.95 (8H, Ar-H),
.10 (1H, t, J = 3.5 MHz, C-H of thiazolidine ring), 3.57
N-(4-((2,4-dioxothiazolidin-5-yl)methyl)phenyl)-4-ethylben-
zamide [TZDs-E] A suspension of 1 g of Intemidiate II in
2
0 mL of 5 % NaOH solution in a stoppered conical flask
(2H, dd, J = 3 MHz, 14 MHz, CH linker), 4.65 (2H, s, CH
2 2
was treated dropwise with a 4-ethylbenzoyl chloride with
constant shaking and cooling in water. The flask was shaken
vigorously for 10 min until the smell of the benzoyl chloride
had disappeared. The solid product was then filtered,
washed with cold water and recrystallized from ethanol
link to benzamide); 13C NMR (DMSO-d6, 125 MHz,):
δ = 179.5 (C, C=O, C-2), 178.1 (C, C=O, C-4), 168.5 (C,
CONH of benzamide), 148.09 (CH, Ar-C, C-1’), 139.4 (CH,
Ar-C, C-4’), 123.0 (2 × CH, Ar-C, C-2’, C-6’), 124.7 (2 ×
CH, Ar-C, C-3’, C-5’), 139.1 (CH, Ar-C, C-1”), 135.1 (CH,
Ar-C, C-4”), 125.8 (2 × CH, Ar-C, C-2”, C-6”), 131.2 (2 ×
CH, Ar-C, C-3”, C-5”), 59.2 (CH, C-5), 45.4 (CH , CH Cl),
(
Yield 86 %); mp 252–256 °C; ultraviolet (UV) (DMSO)
λmax 280; IR (KBr) vmax 3410, 3176, 3057, 1697, 1585,
460, 1315; 1H NMR (DMSO-d6, 500 MHz,): δ = 11.27
1H, s, NH of thiazolidine ring), 9.23 (1H, s, NH of ben-
2
2
1
(
3
5
7.4 (CH , CH linker); Anal. Calcd. for C H ClN O S: C,
2 2 18 15 2 3
7.6; H, 4.0; N, 7.4. Found: C, 56.86; H, 4.13; N, 7.81.
zamide), 7.21–7.92 (8H, Ar-H), 4.01 (1H, t, J = 3.5 MHz,
C-H of thiazolidine ring), 3.54 (2H, dd, J = 3.5 MHz,
N-(4-((2,4-dioxothiazolidin-5-yl)methyl)phenyl)benzamide
[TZDs-B] This compound was prepared from Intemidiate
II and benzoyl chloride following the same procedure
TZDS-E (Yield 88 %) mp 230–234 °C; UV (DMSO)λmax
331; IR (KBr) vmax3446, 3057, 2958, 1732, 1618, 1438;
5
MHz, CH linker), 2.57 (2H, q, J = 3MHz, CH of ethyl
2
2
benzamide), 1.25 (3H, t, J = 2 MHz, CH3 of ethyl
benzamide); 13C NMR (DMSO-d6, 125 MHz,): δ = 175.7
(
C, C=O, C-2), 173.4 (C, C=O, C-4), 167.1 (C, CONH of

Med Chem Res
1
H NMR (DMSO-d6, 500 MHz,): δ = 12.07 (1H, s, NH of
thiazolidine ring), 10.23 (1H, s, NH of benzamide),
.21–7.94 (9H, Ar-H), 4.89 (1H, t, J = 4.5 MHz, C-H of
Acute toxicity test
7
An acute toxicity study was performed for the derivatives
following Organization for Economic Co-operation and
Development (OECD, 2001) guidelines (OECD 425–2001).
Healthy albino rats weighing between 120–150 g were
fasted overnight with water ad-libitum prior to the experi-
ment. On the day of experiment, compounds were admi-
nistered orally at graded doses up to 100 mg/kg in 0.3 %
carboxymethyl cellulose. All the animals were monitored
continuously for 30 min then periodically up to 48 h at 2 h
interval to detect any changes in the autonomic or beha-
vioral responses. They were also checked for tremors,
convulsions, salivation, diarrhea, sleep, lacrimation, heart
rate, pulse rate, blood pressure and feeding behavior as a
sign of acute toxicity.
Q5
thiazolidine ring), 3.10 (2H, dd, J = 9 MHz, 14 MHz, CH2
linker); 13C NMR (DMSO-d6, 125 MHz,): δ = 175.9(C,
C=O, C-2), 171.8 (C, C=O, C-4), 165.5 (C, CONH of
benzamide), 138.09 (CH, Ar-C, C-1’), 134.9 (CH, Ar-C,
C-4’), 120.3 (2 × CH, Ar-C, C-2’, C-6’), 127.4 (2 × CH, Ar-C,
C-3’, C-5’), 131.9 (CH, Ar-C, C-1”), 131.5 (CH, Ar-C,
C-4”), 128.5 (2 × CH, Ar-C, C-2”, C-6”), 129.2 (2 × CH,
Ar-C, C-3”, C-5”), 52.9 (CH, C-5), 36.6 (CH , CH linker);
2
2
Anal. Calcd. for C H N O S: C, 62.5; H, 4.3; N, 8.5.
1
7 14 2 3
Found: C, 61.27; H, 4.19; N, 9.14.
N-(4-((2,4-dioxothiazolidin-5-yl)methyl)phenyl)-4-nitro-
benzamide [TZDs-N] This compound was prepared from
Intemidiate II and 4-nitrobenzoyl chloride following the
same procedure TZDS-E (Yield 82 %) mp 293–297 °C; UV
Experimental design
(
1
DMSO) λmax 364; IR (KBr) vmax 3458, 3049, 2999,
735, 1654, 1539; 1H NMR (DMSO-d6, 500 MHz,):
The whole experimental protocol lasted 14 days. Briefly,
the design consisted of 20 groups of six animals each. All
animals were subjected to induced T2DM, except the
vehicle-treated rats. The T2DM was confirmed by obser-
ving a plasma glucose level of ≥180 mg/dL on the seventh
day of streptozotocin injection. All drugs including the
standard (Pioglitazone) at 1.0-, 10.0- and 100.0 mg/kg dose
levels were administered to their respective groups from day
δ = 11.58 (1H, s, NH of thiazolidine ring), 10.13 (1H, s, NH
of benzamide), 7.11–8.26 (8H, Ar-H), 4.13 (1H, t, J = 4.5
MHz, C-H of thiazolidine ring), 3.59 (2H, dd, J = 7 MHz,
1
=
CONH of benzamide), 154.5 (CH, Ar-C, C-4”), 137.9 (CH,
Ar-C, C-1”),135.07 (CH, Ar-C, C-1’), 133.5 (CH, Ar-C,
C-4’), 122.3 (2 × CH, Ar-C, C-2’, C-6’), 125.4 (2 × CH,
Ar-C, C-3’, C-5’), 129.5 (2 × CH, Ar-C, C-2”, C-6”), 128.2
1 MHz, CH linker); 13C NMR (DMSO-d6, 125 MHz,): δ
2
174.2 (C, C=O, C-2), 168.1 (C, C=O, C-4), 164.5 (C,
8
to 14 of the experimental protocol, except vehicle-treated
T2DM group rats. On days 0 (the day before the strepto-
zotocin was administered), 7 and 14, blood was collected to
measure the level of glucose and triglyceride (TG) in the
animals.
(
2 × CH, Ar-C, C-3”, C-5”), 54.6 (CH, C-5), 34.1 (CH2,
CH linker); Anal. Calcd. for C H N O S: C, 54.9; H,
3
2
17 13 3 5
.5; N, 11.3. Found: C, 55.45; H, 3.80; N, 11.2.
Induction of T2DM
Pharmacological evaluation
The T2DM was induced in the animals using standard
protocol of streptozotocine and nicotinamide administration
Animal
(
Masiello et al., 1998; Garabadu and Krishnamurthy, 2013).
On day 1 of the experimental protocol, streptozotocin
45 mg/kg, intraperitoneally (i.p.); dissolved in 0.1 M of
Male Charles Foster strain albino rats were purchased from
Central Animal House, Institute of Medical Sciences,
Banaras Hindu University and housed in polypropylene
cages under controlled conditions (25 ± 1 °C, 45 %–55 %
relative humidity and 12-h light/dark cycles). The animals
had free access to commercial rat feed and water unless
stated otherwise during the experiment. Animals were
acclimatized for at least 1 week before using them for
experiments and exposed only once to every experiment.
All experiments were conducted in compliance with the
principles of laboratory animal care issued by the National
Institute of Health. Experiments on animals were approved
by the Institutional Animal Ethics Committee of Banaras
Hindu University, Varanasi, India (Protocol number Dean/
(
citrate buffer) and nicotinamide (110 mg/kg, i.p.; dissolved
in physiological saline) were administered at a time lag of
1
5 min.
Estimation of plasma glucose and TG levels
Blood was collected on separate days (0, 7 and 14) from the
retro orbital plexus, and centrifuged (Remu at 3000 × g for
5 min at 4 °C to obtain plasma. The plasma glucose and TG
levels were measured (Muniyappa et al., 2008) in triplicate
using a glucose GOD PAP kit (Priman Instrument Pvt. Ltd.,
India) and TG GPO-PAP kit (Span Diagnostics Ltd., India)
2
014/CAEC/611).

Med Chem Res
Table 1 Anti-hyperglycemic activity on T2DM-induced rats
Table 2 Effect of compounds on T2DM-induced increase in the
plasma triglyceride (mg/dL) levels in rats
Plasma glucose (mg/dL)
Groups
Plasma triglyceride (mg/dL)
Groups
Day-0
Day-7
Day-14
Control
90.5 ± 5.6
Control
77.0 ± 4.8
73.6 ± 1.8
74.9 ± 1.9
75.9 ± 2.3
75.9 ± 2.1
72.4 ± 2.1
72.6 ± 1.5
76.5 ± 5.2
72.6 ± 1.5
74.9 ± 1.9
75.9 ± 2.3
76.5 ± 5.2
72.6 ± 1.5
75.9 ± 2.3
73.6 ± 1.8
75.4 ± 1.9
73.6 ± 1.8
72.6 ± 1.5
75.4 ± 1.9
73.6 ± 1.8
76.7 ± 5.0
486.9 ± 56.3
473.8 ± 43.4
496.1 ± 58.9
503.8 ± 71.9
495.6 ± 48.8
473.3 ± 43.6
476.4 ± 43.4
495.6 ± 48.7
473.3 ± 43.7
476.4 ± 43.4
492.1 ± 52.9
477.2 ± 40.4
510.5 ± 53.5
476.4 ± 43.4
479.7 ± 39.1
494.6 ± 51.1
504.3 ± 73.8
522.1 ± 56.5
504.3 ± 73.8
a
76.5 ± 1.9
512.6 ± 70.1
324.1 ± 12.3
303.8 ± 26.5
a
T2DM
121.1 ± 7.7
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
a
T2DM
a,b
P-1.0
96.2 ± 8.5
a,b
a,b
P-1.0
a,b
P-10.0
94.6 ± 7.9
P-10.0
a,b
P-100.0
93.5 ± 7.5
a,b
P-100.0
290.5 ± 8.8
511.3 ± 56.8
316.1 ± 22.5
a
TZDs-E-1.0
TZDs-E-10.0
TZDs-E-100.0
TZDs-N-1.0
TZDs-N-10.0
TZDs-N-100.0
TZDs-C-1.0
TZDs-C-10.0
TZDs-C-100.0
TZDs-M-1.0
TZDs-M-10.0
TZDs-M-100.0
TZDs-B-1.0
TZDs-B-10.0
TZDs-B-100.0
118.9 ± 12.3
a
TZDs-E-1.0
TZDs-E-10.0
TZDs-E-100.0
TZDs-N-1.0
TZDs-N-10.0
TZDs-N-100.0
TZDs-C-1.0
TZDs-C-10.0
TZDs-C-100.0
TZDs-M-1.0
TZDs-M-10.0
TZDs-M-100.0
TZDs-B-1.0
TZDs-B-10.0
TZDs-B-100.0
a,b
97.3 ± 5.0
a,b
a,b
96.2 ± 5.7
a,b
304.1 ± 6.1
489.6 ± 62.9
463.5 ± 62.2
502.3 ± 81.3
501.1 ± 57.9
268.5 ± 68.0
271.3 ± 60.3
513.3 ± 52.0
505.4 ± 40.5
466.2 ± 62.3
510.3 ± 50.8
502.1 ± 81.3
509.4 ± 50.9
b
a
116.5 ± 5.5
a
a
116.5 ± 5.6
a
a
112.7 ± 6.1
a
a
116.5 ± 5.5
a
a,b
94.9 ± 6.8
a,b
a,b
a
a,b
90.0 ± 7.9
a
115.9 ± 8.1
a
118.6 ± 5.2
a
a
117.3 ± 6.5
a
a
113.8 ± 10.2
a
a
118.6 ± 8.5
a
a
117.3 ± 6.5
a
a
b
All values are mean ± SEM (n = 6).
compared with the control and the T2DM groups, respectively, (one-
way ANOVA followed by Student Newman–Keuls post-hoc test)
P < 0.05 and
P < 0.05
All values are mean ± SEM (n = 6). P < 0.05 and
compared with the control and the T2DM group, respectively (repeated
measures of two-way ANOVA followed by Bonferroni post-hoc test)
P < 0.05
in a spectrophotometer (Beckman Coulter DU 7400
UV–VIS Spectrophotometer, Fullerton, CA).
which involved removing unwanted molecules and other
defects reported by the software. The PPAR γ receptor is a
dimer that has two monomer chains (A and B). For the
purpose of our studies, chain B was deleted and water
molecules near the ligands were retained. Finally, the
energy was minimized and the resulting structure was used
for further docking studies.
Data analysis
All values are shown as the mean ± SEM (n = 6). Plasma
glucose was analyzed using repeated measurements (day 0,
7
and 14) of two-way analysis of variance (ANOVA) fol-
lowing the Bonferroni post-hoc test. The level of plasma TG
was analyzed on day 14 of the experimental protocol using
one-way ANOVA followed by the Student Newman–Keuls
post-hoc test. A level of P < 0.05 was considered as sig-
nificant in the whole data analysis.
Preparation of ligand
Designed molecules whose docking studies were performed
were refined using ligPrep module. The minimized energy
three-dimensional structure of molecule was obtained using
Optimized Potential for Liquid Simulations-2005 force field.
Docking
Docking studies
Docking studies were performed using Schrodinger 9.0
maestro version on PPAR γ.
Docking was carried out using the ‘standard precision’
Glide algorithm in Schrodinger. Extra precision and write
XP descriptor information was used to generate favorable
ligand poses that were further screened through filters to
examine the spatial fit of the ligand in the active site. Ligand
poses that passed the initial screening were subjected to
evaluation and minimization of grid approximation.
Selection and preparation of the protein
The crystal structure of PPAR-γ was obtained from the
protein data bank (pdb code: 3CSB). The protein was pre-
pared using protein preparation wizard in maestro 9.3,

Med Chem Res
Fig. 2 Binding interaction
between PPAR- γ with
compound TZDs-Z a and
standard Pioglitazone b

Med Chem Res
Table 3 Docking score and binding energy of synthesized
compounds
significantly higher glucose level in the T2DM group ani-
mals than the control rats on day 7, which was maintained
up to day 14. However, pioglitazone (P; 1.0, 10.0 and 100.0
mg/kg), TZDs-E (10.0 and 100.0 mg/kg) and TZDs-C (10.0
and 100.0 mg/kg) had attenuated T2DM-induced increase in
the plasma glucose level by day 14.
S. no
Compound
G-score
G energy
1
2
3
Pioglitazone
TZDs-E
−7.75
−6.38
−6.14
−41.45
−36.22
−35.89
TZDs-C
Table 2 shows the effect of the synthesized compounds
on the T2DM-induced changes in the plasma TG level in
the rats. Statistical analysis by one-way ANOVA revealed
that there were significant differences among groups (F (19,
100) = 12.67; P < 0.05). Post-hoc study showed that there
was a significantly higher TG level in the T2DM group
animals than in the control rats. Moreover, P (1.0, 10.0 and
100.0 mg/kg), TZDs-E (10.0 and 100.0 mg/kg) and TZDs-C
Scoring was then carried on energy-minimized poses to
generate a Glide score.
Results and discussion
Chemistry
(10.0 and 100.0 mg/kg) attenuated the T2DM-induced
increase in the plasma glucose level.
All the proposed novel thiazolidine-2,4-diones were suc-
cessfully synthesized and confirmed through spectral char-
acterization and elemental analysis. FT-IR spectra showed
characteristic amide (C=O) stretching between 1674 and
Docking
Docking studies were conducted to get a comprehensive
insight about the nature of interaction between the com-
pounds and target Fig. 2. Pioglitazone was taken as the
reference compound. Glide score was predicted by Impact
version v57111. Table 3 shows the binding energies of
the synthesized compound and pioglitazone, Fig. 2 and
Fig. 3 show the type of interaction between compounds and
target. The synthesized compounds showed both, hydrogen
bonding and hydrophobic interaction with the target. The
compounds show interactions with amino acid residue
Phe282, Cys285, Hie449, Ser289, Lys 261, Glu 259.
−1
1
3
628 cm and amide (N-H) stretching between 3458 and
−1
410 cm , confirming the formation of the amide linkage.
−1
Peaks between 3057 and 2924 cm were observed for the
alkyl (C-H) stretching, which indicates the presence of a
methylene bridge.
1
H NMR spectra showed a characteristic singlet peak
between 8.01 and 8.34 ppm for the amide proton (CONH),
and several multiplets were observed between 6.78 and
8
.09 ppm, representing the aromatic protons.
Acute oral toxicity study
Discussion
All the synthesized compounds were assessed for acute oral
toxicity profile in rats before the anti-diabetic study (OECD
The present study describes the successful synthesis of novel
derivatives of pioglitazone that exhibit anti-hyperglycemic
and anti-hyperlipidemic effects in T2DM animals. PPAR
gamma is a nuclear receptor that regulates carbohydrate and
lipid metabolism, and attenuates peripheral insulin resistance
through reducing blood glucose and TG levels in T2DM
subjects (Dumasia et al., 2005). In the present work, thia-
zolidinedione was chosen as the central nucleus as it is
reported to interact with PPAR gamma receptors. Analogs of
pioglitazone were designed with an amide linkage instead of
an ether linkage, and their structures were confirmed by
spectral and elemental data analysis. In the oral acute toxi-
city study, the screened molecules were well tolerated for
14 days. Furthermore, in the anti-diabetic study, compounds
TZDs-C and TZDs-E showed significant anti-hyperglycemic
and anti-hyperlipidemic effects in the T2DM animals. This
indicates that these molecules are promising insulin sensiti-
zers, and may exert their therapeutic activity through PPAR
gamma receptors. Molecular docking studies were carried
4
25–2001). All the compounds were well tolerated up to a
dose of 500 mg/kg. In addition, no mortality was observed
up to 14 days of post administration, even at the highest
dose of 500 mg/kg, which was considered to be an appre-
ciable margin of safety.
Anti-hyperglycemic and anti-hyperlipidemic activity
The synthesized compounds were evaluated for their anti-
hyperglycemic and anti-hyperlipidemic activity in T2DM
rats. Table 1 shows the effect of the synthesized compounds
on the T2DM-induced changes in the plasma glucose levels
in the rats. Statistical analysis by repeated measures of two-
way ANOVA revealed that there were significant differ-
ences in the plasma glucose level among groups (F (19,
2
40) = 7.16; P < 0.05) and times (F (2, 240) = 495.0;
P < 0.05), and also between groups and times (F (38, 240)
3.18; P < 0.05). Post-hoc study showed that there was
=

Med Chem Res
Fig. 3 Hydrophobic interaction
of TZDs-E a and Pioglitazone b
with PPAR-γ

Med Chem Res
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Technology (BHU) for providing financial assistance through
Sprouting grant for faculty (SGF) and Institute research project (IRP)
scheme.
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