Design and synthesis of 2,4-disubstituted polyhydroquinolines as prospective antihyperglycemic and lipid modulating agents

Article abstract of DOI:10.1016/j.bmc.2009.11.061

A series of 2,4-disubstituted polyhydroquinoline were synthesized and evaluated for their in vivo antihyperglycemic as well as antidyslipidemic activities. Several synthesized compounds have exhibited promising in vivo antihyperglycemic in SLM, STZ-S, and db/db mice model along with significant lipid and TG modulating activity. All these compounds were evaluated in various in vitro models of diabetes to know the possible mechanism of their antihyperglycemic action. Interestingly, compounds 3a-r (diaryl substitution) have exhibited promising protein-tyrosine phosphatase 1B (PTP1B) inhibitory activity whereas, compounds 5a-d (acid substituted) have shown significant glycogen phosphorylase activity.

Full text of DOI:10.1016/j.bmc.2009.11.061

Bioorganic & Medicinal Chemistry 18 (2010) 4138–4148
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Design and synthesis of 2,4-disubstituted polyhydroquinolines as prospective
antihyperglycemic and lipid modulating agents
a
a
a
Atul Kumar a, , Siddharth Sharma , Vishwa Deepak Tripathi , Ram Awatar Maurya ,
*
Swayam Prakash Srivastava b, Gitika Bhatia b, A. K. Tamrakar b, Arvind Kumar Srivastava b
a Medicinal and Process Chemistry Division, Central Drug Research Institute, Council of Scientific and Industrial Research, Lucknow 226 001, India
b Biochemistry Division, Central Drug Research Institute, Council of Scientific and Industrial Research, Lucknow 226 001, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of 2,4-disubstituted polyhydroquinoline were synthesized and evaluated for their in vivo antihy-
perglycemic as well as antidyslipidemic activities. Several synthesized compounds have exhibited prom-
ising in vivo antihyperglycemic in SLM, STZ-S, and db/db mice model along with significant lipid and TG
modulating activity. All these compounds were evaluated in various in vitro models of diabetes to know
the possible mechanism of their antihyperglycemic action. Interestingly, compounds 3a–r (diaryl substi-
tution) have exhibited promising protein-tyrosine phosphatase 1B (PTP1B) inhibitory activity whereas,
compounds 5a–d (acid substituted) have shown significant glycogen phosphorylase activity.
Ó 2009 Published by Elsevier Ltd.
Received 23 October 2009
Revised 25 November 2009
Accepted 26 November 2009
Available online 6 December 2009
Keywords:
Polyhydroquinoline
Antihyperglcemic
Antidyslipidemic
PTP-1B
Glycogen phosphorylase
1. Introduction
The discovery of new lead structure is important area of research
in medicinal chemistry as new chemical entity is continue to fall and
Recently, diabetes has again focused the world’s attention as
approximately 330 million will be afflicted worldwide by 2025
with a 122% overall increase.1 Non-insulin dependent diabetes
mellitus (Type-2 diabetes), is a complex metabolic disorder of
heterogeneous etiology and is primarily characterized by insulin
resistance and abnormal insulin secretion. These metabolic disor-
ders cause hyperglycemia,2,3 which is significantly responsible for
diabetic complications. Complication of diabetes4 includes neurop-
athy, nephropathy, retinopathy, obesity, dyslipidemia, hyperten-
sion, and other cardiovascular diseases and these are believed to
be triggered by excessive protein glycation, which is due to higher
levels of circulating glucose (hyperglycemia).
The increased risk of cardiovascular disease in Type 2 diabetes
is commonly due to abnormalities in the quantity as well as quality
of plasma lipoproteins. Besides this elevated triglyceride and high
density lipoprotein cholesterol, diabetic dyslipidemia is also asso-
ciated with predominance of low density lipoproteins.
toxicity of the present drugs becomes a dilemma. Among various
biologically active heterocyclic scaffolds, Hantzsch 1,4-dihydropyri-
dines (1,4-DHPs) are an important class of biologically active hetero-
cycles. The prominent biological activities associated with 1,4-
dihydropyridines are as Ca2+ channel blockers (drugs for the treat-
ment of cardiovascular diseases)9 and hypertension (nifedipine,
nicardipine, amlodipine, and felodipine). Beside, this dihydropyri-
dine skeleton is common in many biological activities such as anti-
hypertension,10 antioxidant,11 antiviral,12 and anticancer activity.13
In continuation to our antidiabetic drug discovery program,14
we have designed 2,4-disubstituted polyhydroquinoline deriva-
tives based on the some recent reports that, indicate 1,4-DHPs
may act as a lead in the antidiabetic drug discovery, such as glyco-
gen phosphorylase inhibitors (215, 3,16 4,16), cerivastatin 5 (lipid
lowering agent), and 617 for the treatment of metabolic disorder
(Fig. 1). A series of 2,4-disubstituted polyhydroquinoline deriva-
tives have been synthesised and evaluated for their antidiabetic
and antidyslipidemic activity in various in vivo and in vitro models.
Dyslipidemia is connected with insulin insensitivity and associ-
ated to increased atherosclerosis susceptibility.4–8 Therefore, it is
win to win state for antidiabetic therapy, if any chemical entity
takes care of diabetes as well as dyslipidemia.
2. Results and discussion
2.1. Chemistry
The different chalcones were synthesized by the previously
reported method of Sogawa et al.18 2,4-Disubstituted polyhydro-
quinoline, were synthesized via a three-component reaction of
* Corresponding author. Tel.: +91 522 2612411; fax: +91 522 2623405.
E-mail address: dratulsax@gmail.com (A. Kumar).
0968-0896/$ - see front matter Ó 2009 Published by Elsevier Ltd.
doi:10.1016/j.bmc.2009.11.061

A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
4139
F
F
Cl
O
O
O
O
OH
OH
HO
HO
N
N
N
2
4
3
(Antidiabetic)
CN
N
F
HN
(CH2)3
HN
)
c
i
O
m
OH OH
O
e
N
d
O
O
i
p
i
l
O
OH
R1
s
O
R3
O
N
H
y
R1
d
i
N
CH2Ph
t
n
N
1
A
H
(
R3 = Ph/ COOEt /COOH
5
6
Antihypertensive
Cl
Cl
COOEt
Cl
CH2OOEt
MeOOC
NO2
MeOOC
MeOOC
COOMe
O
N
H
NH2
N
H
N
H
8
7
9
Figure 1. Design of 2,4-disubstituted polyhydroquinoline based on dihydropyridine scaffolds showing antidiabetic, antihypertensive and dyslipidemic activity.
Table 1
substituted 1,3-cyclicdiketone, chalcones, and ammonium acetate
Synthesis of 2,4-diaryl polyhydroquinoline via a multi-component reactiona
(Scheme 1). Number of acidic catalysts used such as boric acid, so-
R1
R2
R3
Time (h)
Productb
dium dodecyl sulfate (SDS), camphor sulfonic acid, para toluene
sulfonic acid (PTSA), and tetra butyl ammonium iodide (TBAI).
PTSA found to be the best among all the catalysts used. Although
the reaction was sluggish in various solvents, MeOH was found
to be the best among all the solvents used for the reaction. Chal-
cones (1 mmol), 5,5-dimethyl-1,3-cyclohexanedione (1 mmol),
ammonium acetate (2 mmol) were refluxed in methanol in the
presence of PTSA to afford desired product in excellent yields. All
the results are summarised in Table 1.
Entry
1
2
3
4
5
6
7
8
9
10
11
12
13
14
CH3
CH3
CH3
CH3
CH3
CH3
H
H
H
CH3
CH3
H
Ph
Ph
Ph
Ph
Ph
Ph
4-OMe Ph
Ph
Ph
Ph
4-CH3 Ph
Ph
4-CH3 Ph
Ph
Ph
2.5
1.5
1.7
2.3
2.7
2.9
2.4
1.7
1.5
2.6
3.7
2.3
3.0
2.5
3a
3b
3c
3d
3e
3f
3g
3h
3i
3j
3k
3l
3m
3n
4-NO2 Ph
4-Cl Ph
4-F Ph
4-OMe Ph
Ph
Ph
4-Cl Ph
4-F Ph
Ph
Indolyl
Ph
b,c-Unsaturated a-ketoester (2o–s) were synthesized in two
steps procedure. Firstly, aryl aldehyde and pyruvic acid were con-
densed in the presence of KOH to give potassium (E)-2-oxo-4-phe-
nylbut-3-enoates. Esterification of potassium (E)-2-oxo-4-
phenylbut-3-enoates gave (E)-ethyl 2-oxo-4-phenylbut-3-enoates
(2o–s) in excellent yields (Scheme 2).
Some newer ethyl 7,7-dimethyl-5-oxo-4-phenyl-1,4,5,6,7,8-
hexahydroquinoline-2-carboxylate derivatives 4o–s (Scheme 3.)
have been synthesized for the further diversification in our polyhy-
H
CH3
4-OMe Ph
Furyl
a
Experimental conditions: all the reactions were performed at refluxing in PTSA
in 10 mol %.
b
Product characterized by spectroscopic (1H and 13C NMR) data.
droquinoline core motif to generate ester group at C-2 position via
a two step protocol. Reaction of dimedone, and b, -unsaturated a-
ketoester in presence of acidic catalyst gave corresponding lactol
derivatives (3o–s) in excellent yields. These lactol derivatives were
transformed in to polyhydroquinoline derivatives by the reaction
with ammonium acetate in quantitative yield.
Previously, it have been reported that several antidiabetic
agents contain carboxylic acid group as key pharmcophores17
(Fig. 1). Keeping this hypothesis in mind, we have hydrolyzed the
ester group present at the C-2 position to generate corresponding
carboxylic acid group (5a–e) (Scheme 4).
c
O
O
R2
R2
O
PTSA/MeOH
Reflux
R1
R1
R1
R1
O
N
H
R3
R3
NH4OAC
1a-1b
1a: R1 = H, 1b: R1 = CH3
Scheme 1. Synthesis of 2,4-diaryl polyhydroquinolines.
3a-3n
2a-2n

4140
A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
O
O
O
O
a
OH
b
OK
OEt
Ar
H
Ar
Ar
O
O
O
Ar = C6H5 (2o)
4-MeO-C6H4
4-Cl-C6H4 (2q)
2r
2p
º
(
)
(a) KOH, MeOH, RT, 10-15 h; (b) AcCl, EtOH, 70 C, 6-8 h.
4-F-C6H4
(
)
2s
(
4-Br-C6H4
)
Scheme 2. Synthesis of (E)-ethyl 2-oxo-4-arylbut-3-enoates.
O
R2
O
R2
O
R2
PTSA
NH4OAc
OH
R3
MeOH, RT
O
R3
EtOH, Reflux
O
O
R3
N
H
R3 = COOEt
2o-2s
4o-4s
3o-3s
Scheme 3. Synthesis of substituted lactol and polyhydroquinoline derivatives.
R2
Recently, we have developed Human hemoglobin (HbA) as an
efficient catalyst for the oxidative aromatization of 1,4-dihydro-
pyridines (1,4-DHPs) and pyrazolines with hydrogen peroxide in
phosphate buffer.19 In order to evaluate the effect of aromatization
on antidiabetic activity of 2,4-disubstituted polyhydroquinoline
were aromatized to corresponding tetrahydroquinoline ring 6a–c
in excellent yields using Human hemoglobin (HbA)/H2O2 catalytic
system (Scheme 5).
O
R2
N
O
HbA (0.1 mmol), 30% (w/v) H2O2 (2 mmol)
Phosphate buffer (pH = 6.5) rt, Stirr
R1
R3
N
H
R1
R1
R3
R1
3d, 3h, 3m
6a, 6b, 6c
Scheme 5. Aromatization of polyhydroquinoline with HbA/H2O2 catalyst system.
glucose-6-phosphatase,20 glycogen phosphorylase,20
protein-tyrosine phosphatase 1B and dipeptidyl peptidase IV (DPP
IV) enzymes at 100 M concentration (Table 4) to evaluated possible
a-glucosidase,
2.2. Biological activity
l
2.2.1. In vivo antihyperglycemic activity
mechanism of action for compounds exhibiting significant antidia-
betic activity in in vivo models.
All the synthesized compounds (3a–s), (4o–s), (5a–e), and (6a–
c) were evaluated for their antihyperglycemic activity in sucrose-
loaded model (SLM) and streptozotocin (STZ-S) induced b-cell
damaged diabetic model of Sprague–Dawley strain male albino
rats.
Compounds 3d (41.3%), 3i (34.5%), 4r (35.9%), and 5b (42.7%)
have shown promising blood glucose lowering activity in SLM
model. Compounds showing blood glucose lowering activity were
further tested for antidiabetic activity in sucrose-challenged strep-
tozotocin (STZ-S) induced diabetic rat model. The compounds 3d
(32.3%), 3i (24.1%), 4r (23.4%), 5b (33.2%) exhibited comparable
or higher antihyperglycemic activities to standard drugs, metfor-
min (20.2%), and glybenclamide (28.4%) in STZ model after 24 h
treatment. All the results are summarised in Table 2.
Compounds (3d, 5b) exhibited 13.2% (3d), 17.9% (5b) after
5 days and 32.6% (3d), and 39.6% (5b) blood glucose lowering
activity after 10 days treatment. The standard drug rosiglitazone
showed 22.6% after 5 days and 52.9% blood glucose lowering activ-
ity after 10 day treatment in similar conditions at a dose of
100 mg/kg (Table 3).
The effect of compounds on glucose-6-phosphatase was studied
by the method of Hubscher and West.21 The dipeptidyl peptidase
enzyme inhibition assays were performed according to the meth-
ods of Wright et al.22 The glycogen phosphorylase activity is mea-
sured by the modified method of Rall et al.23 The
a-glucosidase
activity was determined by the method of Lebovitz et al.24 Pro-
tein-tyrosine phosphatase 1B inhibitory activity was determined
by the modified method25 using pNPP as substrate. Compounds
(3a–s) were showing poor inhibitory activities for glucose-6-phos-
phatase, glycogen phosphorylase,
a-glucosidase, and dipeptidyl
peptidase IV (DPP IV) enzymes, whereas compounds ( 3b–3e, 3i,
3n, 4r, and 4s) have shown promising PTP 1B inhibitory activity.
All the compounds with promising in vivo activity showed good
PTP 1B inhibitory activity. Compound 3n exhibited excellent PTP
1B inhibitory activity but found inactive in in vivo model. IC50
value of active compounds concluded that protein-tyrosine phos-
phatase 1B (PTP1B) may be a possible target for these antihyper-
glycemic compounds. Very interesting in vitro results were
obtained in case of 5a–e, as 5b was the most active in vivo
compound of the series and found to inhibit the glycogen phos-
phorylase more significantly instead of PTP 1B. Exact reason was
not elucidated but –COOH group in 5a–e may be playing important
2.2.2. In vitro antihyperglycemic activity
All the synthesized compounds (3a–s), (4o–s), (5a–e), and (6a–c)
were evaluated for in vitro antihyperglycemic activity against
R2
O
R2
O
NaOH
MeOH
N
H
COOH
N
H
4r
COOEt
, R2 = Ph
2
2
5a
5b
5c
, R2 = Ph
, R2 = 4-OMe Ph
, R2 = 4-Cl Ph
5d
5e
4o
4p
4q
, R = 4-Cl Ph
,
R2 = 4-F Ph
, R2 = 4-Br Ph
,
R2 = 4-Br Ph
, R = 4-OMe Ph
, R2 = 4-Cl Ph
4s
Scheme 4. Hydrolysis of ester group of polyhydroquinolines.

A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
4141
Table 2
In vivo antihyperglycemic activity profile of compounds in SLM and STZ-S modelsa
c
Entry
R1
R2
R3
Compounds
% Blood glucose lowering activityb,
STZ-S
SLM
d
5 h
24 h
ED50
1
2
3
4
5
6
7
8
9
CH3
CH3
CH3
CH3
CH3
CH3
H
H
H
CH3
CH3
H
Ph
Ph
Ph
Ph
Ph
Ph
4-OMe Ph
Ph
Ph
Ph
4-CH3 Ph
Ph
4-CH3 Ph
Ph
3a
3b
3c
3d
3e
3f
3g
3h
3i
3j
3k
3l
3m
3n
3o
3p
3q
3r
3s
4o
4p
4q
4r
4s
5a
5b
5c
5d
5e
6a
6b
6c
24.9
14.1
23.5
41.3
17.8
17.3
19.7
24.7
34.5**
11.4
17.3
13.4
29.6
14.6
NA
17.3
ND
18.9
29.9
8.8
11.9
15.1
12.9
19.6**
ND
8.1
ND
17.5
ND
ND
ND
ND
ND
ND
7.1
ND
ND
24.8**
ND
13.3
ND
4-NO2 Ph
4-Cl Ph
4-F Ph
4-OMe Ph
Ph
16.4
32.3
10.9
11.7
13.8
16.9
24.1**
ND
13.2
ND
19.4
ND
ND
ND
ND
ND
ND
11.3
ND
ND
23.4**
ND
16.7
33.2
12.2
23.6
13.6
ND
**
***
***
47.3
Ph
4-Cl Ph
4-F Ph
Ph
Indolyl
Ph
4-OMe Ph
Furyl
Ph
4-OMe Ph
4-Cl Ph
4-F Ph
4-Br Ph
Ph
4-OMe Ph
4-Cl Ph
4-F Ph
4-Br Ph
Ph
4-OMe Ph
4-Cl Ph
4-F Ph
4-Br Ph
4-F Ph
4-Cl Ph
4-OMe Ph
—
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
H
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
H
Ph
COOEt
COOEt
COOEt
COOEt
COOEt
COOEt
COOEt
COOEt
COOEt
COOEt
COOH
COOH
COOH
COOH
COOH
Ph
NA
NA
15.3
NA
16.6
12.9
11.4
35.9**
11.4
28.9
42.7
13.7
38.7
31.7
NA
18.3
**
***
**
**
29.7
16.6
27.8
17.8
ND
48.6
**
***
Ph
Ph
—
—
NA
NA
13.5
32.5
ND
ND
—
—
Metformin
Glybenclamide
18.0
25.6
20.2
28.4
a
b
c
Dose = 100 mg/kg.
NA = not active, ND = not determine.
*p <0.05, **p <0.01 and ***p <0.001 versus vehicle treated control and Statistical analysis was made by Dunnett’s test (Prism Software).
d
mg/kg.
Table 3
Antihyperglycemic activity of 3d and 5b in db/db mice modelc
Entry
Compounds
na
5 dayb
10 dayb
32,480 5098
1
2
3
4
Control
3d
5b
5
5
5
5
30,730 3687
26,670 3058 (ꢀ13.2)*
21,870 3069 (ꢀ32.6)**
25,250 2220 (ꢀ17.9)*
19,630 2467 (ꢀ39.6)**
**
***
Rosiglitazone
23,780 2261 (ꢀ22.6)
15,280 2300 (ꢀ52.9)
a
b
c
No. of mice.
Mean of glucose (AUC) values SD. *p<0.05, **p <0.01 and ***p <0.001 versus vehicle.
% Lowering values are given in parentheses.
role in reversal the mode of action. Further optimization is also
going on in our group to increase the glycogen phosphorylase
inhibitory activity. None of the aromatized 2,4-disubstituted poly-
hydroquinoline 6a–c found to inhibit any of the performed in vitro
assays.
a dose of 100 mg/kg, which showed a decrease in plasma levels of
TC, PL and TG by 37.5%, 39.4%, and 36.2%, respectively (Table 5).
The SAR activity revealed interesting finding that 2,4-diaryl
substituted compounds were exhibiting antidiabetic activity via
inhibition of PTP-1B, whereas 2-carboxylic acid substituted com-
pounds (5) were showing promising glycogen phosphorylase
inhibitory activity. It is noteworthy to report that compounds
having ester group at position-2- were exhibiting moderate PTP-
1B and glycogen phosphorylase inhibition along with significant
in vivo antidiabetic activity. Aromatization of dihydropyridine ring
(6) resulted in complete loss of antidiabetic activity (Fig. 2). These
finding indicates that dihydropyridine ring is important for hyper-
glycemic activity.
2.2.3. In vivo antidyslipidemic activity
Compounds showing significant in vivo antidiabetic activity
were further evaluated for their antidyslipidemic activities in a tri-
ton model. Compound 3c, 3d, and 5b showed significant lipid low-
ering activity in plasma level of TC, PL, and TG, respectively, while
other compounds showed moderate lipid lowering activity as com-
pared to triton. All the activity were compared with Gemfibrozil at

4142
A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
Table 4
In vitro antihyperglycemic activity of (3a–s), (4o–s), (5a–e), and (6a–c)
Compounds
% Inhibitiona
Glycogen phosphorylase
Glucose-6-phosphatase
a-Glucosidase
DPP IV
PTP-1B
3a
3b
3c
3d
3e
3f
3g
3h
3i
3j
3k
3l
3m
3n
3o
NI
NI
NI
NI
5.89
14.58
NI
18.41
21.83
19.34
17.43
31.67
39.67
NI
13.81
18.88
NI
34.32
22.35
12.13
NI
NI
22.4
NI
27.9
NI
NI
12.0
NI
NI
NI
8.6
NI
NI
NI
NI
NI
NI
18.8
NI
18.6
NI
41.8
84.7 (6.1)
84.9 (6.3)
88.2 (2.9)
71.8 (14.5)
53.4
51.2
24.7
70.3 (12.6)
62.3 (19.6)
54.5
7.16
7.38
NI
8.34
NI
8.21
21.89
NI
26.71
27.66
17.28
11.38
NI
NI
8.3
7.3
NI
33.3
56.8
82.5 (7.7)
21.3
NI
3p
NI
NI
NI
NI
13.6
3q
NI
7.6
NI
NI
16.5
3r
NI
14.9
NI
NI
5.3
3s
NI
12.3
NI
NI
4.7
4o
4p
4q
4r
4s
5a
5b
5c
5d
5e
6a
6b
NI
49.45
40.39
56.71
61.23
49.3
77.3 (6.1)
78.3 (5.9)
63.4 (9.7)
46.6
81.3 (5.3)
20.3
13.8
31.1
—
NI
NI
NI
15.8
NI
4.6
11.2
NI
1.4
NI
NI
43.3
66.7
53.8
87.2 (4.7)
65.6 (16.6)
28.9
35.4
25.4
27.0
12.3
7.9
18.6
23.81
11.27
NI
NI
17.1
NI
NI
16.8
NI
11.1
11.3
NI
16.47
12.5
9.8
NI
14.6
5.7
NI
NI
NI
13.1
29.4
22.0
57.8
NI
NI
NI
—
6c
9.1
Sod. ortho-vanadate
—
a
Compounds were evaluated at 100
lM concentration; NI means no inhibition. IC50 (lM) values are given in parentheses.
Table 5
In vivo antidyslipidemic activities of selected compoundsc
Entry
Compounds
nb
TCa
225 12.85
PLa
TGa
310 12.40
1
2
3
4
5
6
7
8
9
Control
3a
3c
3d
3h
3r
5a
5b
6
6
6
6
6
6
6
6
6
6
227.2 10.7
184.74 10.2 (ꢀ17.89)*
167.49 9.7 (ꢀ25.56)**
174.48 10.6 (ꢀ22.45)
209.92 11.8 (ꢀ6.7)
183.34 7.2 (ꢀ19.23)*
259.0 13.2 (ꢀ16.45)*
175.99 8.7 (ꢀ22.47)**
250.10 14.7 (ꢀ19.32)**
**
*
**
184.25 7.6 (ꢀ18.83)
213.60 6.8 (ꢀ5.9)
228.34 14.6 (ꢀ26.34)
285.54 15.8 (ꢀ7.89)
284.58 16.4 (ꢀ8.2)
*
*
201.82 12.4 (ꢀ10.3)
187.95 6.4 (ꢀ17.2)
*
204.77 11.3 (ꢀ8.99)
169.59 9.8 (ꢀ24.63)**
185.73 8.7 (ꢀ17.45)*
140.49 7.4 (ꢀ37.56)**
209.08 7.1 (ꢀ7.89)
267.03 14.6 (ꢀ13.86)
177.46 8.8 (ꢀ21.82)*
192.08 9.7 (ꢀ15.38)*
137.49 6.4 (ꢀ39.43)**
250.04 13.8 (ꢀ19.34)**
272.89 16.7 (ꢀ11.97)*
197.74 10.4 (ꢀ36.21)**
5c
10
Gemfibrozil
TC, total cholesterol; PL, phospholipid; TG, triglyceride. Values are mean SD, *p <0.05, **p <0.01 and ***p <0.001 versus vehicle.
a
mg/dL.
n = no. of animals.
% Lowering values are given in parentheses.
b
c
more significantly than PTP-1B and our studies shows that the
acidic group is beneficial for glycogen phosphorylase inhibitory
activity. Compound 3c, 3d, and 5b were showing promising antidy-
slipidemic activity in triton induced rat model. Thus, 2,4-disubsti-
tuted polyhydroquinoline show remarkable promise for further
study as antidiabetic candidates.
3. Conclusion
In conclusion, we have synthesized a series of 2,4-disubstituted
polyhydroquinoline and evaluated them for their antihyperglyce-
mic as well as antidyslipidemic activities. Some compounds of
the series (3d, 3i, 5b, and 5d) have been exhibiting antihyperglyce-
mic activity comparable to standard drugs along with significant
lipid and TG lowering activity. Interestingly, in vitro antihypergly-
cemic activity evaluation exhibited that compounds 3d (diaryl
4. Experimental
4.1. Chemistry
substituted) (IC50 = 2.9 lM) and 4r (IC50 = 4.7 lM) are potential
PTP-1B inhibitors thereby revealing their possible mechanism of
antidiabetic action. Whereas, compounds 5a–e containing carbox-
ylic group (–COOH) group inhibits the glycogen phosphorylase
Unless otherwise specified all the reagents and catalysts were
purchased from Sigma–Aldrich and were used without further

A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
4143
4.1.1.2. (E)-Ethyl
4-(4-methoxyphenyl)-2-oxobut-3-enoate
EWG (Flouro group) enhance the
anti-hyperglycemic activity
(2p). Yield (92.56%), ESIMS (m/z) = 235 (M+H)+. 1H NMR (CDCl3,
300 MHz) d: 1.41 (t, J = 7.2 Hz, 3H), 3.86 (s, 3H), 4.39 (q,
J = 7.2 Hz, 2H), 6.93–7.61 (m, 4H), 7.25 (d, J = 16.0 Hz, 1H), 7.84
(d, J = 16.0 Hz, 1H). 13C NMR (CDCl3, 75 MHz) d: 14.2, 55.6, 62.5,
114.8, 118.4, 126.9, 131.2, 148.4, 162.7, 162.8, 182.9.
R
G
=
l
y
C
p
c
O
h
o
i
n
g
O
h
en
H
i
b
,
i
t
e
o
n
r
o
h
y
s
an
p
F
act
h
ce
or
i
v
y
t
4.1.1.3. (E)-Ethyl 4-(4-chlorophenyl)-2-oxobut-3-enoate (2q). Yield
(87.38%), ESIMS (m/z) = 238 (M+H)+. 1H NMR (CDCl3, 300 MHz) d: 1.42
(t, J = 7.2 Hz, 3H), 4.40 (q, J = 7.2 Hz, 2H), 7.35 (d, J = 16.0 Hz, 1H), 7.39–
7.58 (m, 4H), 7.81 (d, J = 16.0 Hz, 1H). 13C NMR (CDCl3, 75 MHz) d: 14.0,
62.5, 120.8, 129.3, 130.1, 132.4, 137.5, 146.6, 161.9, 182.4.
i
h
l
t
a
y
e
3
s
e
O
N
i
a
t
n
r
o
t
i
g
d
e
i
N
R3
a
n
b
4.1.1.4. (E)-Ethyl 4-(4-fluorophenyl)-2-oxobut-3-enoate (2r). Yield
(85.50%), ESIMS (m/z) = 223 (M+H)+. 1H NMR (CDCl3, 300 MHz) d: 1.42
(t, J = 7.2 Hz, 3H), 4.40 (q, J = 7.2 Hz, 2H), 7.10–7.67 (m, 5H), 7.84 (d,
J = 16.0 Hz, 1H). 13C NMR (CDCl3, 75 MHz) d:14.0, 62.5, 120.8, 129.3,
130.1, 132.4, 137.5, 146.6, 161.9, 182.4.
i
e
s
t
e
i
c
s
s
t
a
e
c
n
t
i
t
i
v
a
i
l
y
f
o
r
R3= COOEt exhibiting
mixed respones
4.1.1.5. (E)-Ethyl 4-(4-bromophenyl)-2-oxobut-3-enoate (2s). Yield
(88.89%), ESIMS (m/z) = 282, 284 (M+H)+. 1H NMR (CDCl3, 300 MHz) d:
1.42 (t, J = 7.2 Hz, 3H), 4.40 (q, J = 7.2 Hz, 2H), 7.37 (d, J = 16.0 Hz, 1H),
7.49–7.58 (m, 4H), 7.79 (d, J = 16.0 Hz, 1H). 13C NMR (CDCl3, 75 MHz) d:
13.7, 62.2, 120.7, 125.7, 130.0, 132.0, 132.6, 146.3, 161.6, 182.1.
Figure 2. Structure–activity relationship and pharmacophore development of
polyhydroquinoline.
any purification. The common solvents were purchased from Ranb-
axy. Organic solutions were concentrated under reduced pressure
on a Büchi rotary evaporator. Chromatographic purification of
products was accomplished using flash chromatography on 230–
400 mesh silica gel. Reactions were monitored by thin-layer chro-
matography (TLC) on 0.25 mm silica gel plates visualized under UV
light, iodine or KMnO4 staining. 1H and 13C NMR spectra were re-
corded on a Brucker DRX-300 Spectrometer. Chemical shifts (d)
are given in ppm relative to TMS and coupling constants (J) in
Hz. IR spectra were recorded on a FT IR spectrophotometer Shima-
dzu 8201 PC and are reported in terms of frequency of absorption
(cmꢀ1). Mass spectra (ESIMS) were obtained by Micromass Quattro
II instrument.
4.1.2. Typical experimental procedure for the synthesis of 2,4-
Substituted polyhydroquinoline (3a–n)27
Mixture of 5,5-dimethyl-cyclohexane-1,3-dione (1.2 mmol) and
1,3-diphenyl-propenone (1 mmol), ammonium acetate (2 mmol),
PTSA (20 mol %) were taken in 50 mL round bottem flask, was
added to MeOH. The reaction mixture was stirred at 80 °C for
2.5 h. After completion of the reaction as indicated by TLC, then
treated with EtOAc followed by water and brine solution and dried
with anhydrous Na2SO4. The solution was concentrated in vacuum
to give the crude solid. The crude solid was purified by recrystalli-
zation from methanol. Although in case of nitro substituent, crude
was purified by column chromatography with EtOH/hexane 1:5 as
elutant. Compounds 3b–n were prepared from the corresponding
chalcone following similar procedure.
4.1.1. Typical experimental procedure for the synthesis of b,
c-
unsaturated -ketoester (2o-s)
a
To a benzaldehyde (10.6 g, 0.1 mol) solution in methanol
(15 mL) was added pyruvic acid (8.6 g, 0.10 mmol), and the mix-
ture was cooled to 10 °C under the atmosphere of nitrogen. To this
was added a solution of KOH (8.4 g, 0.15 mmol) in methanol drop-
wisely at 15–20 °C, after the addition. Then the ice-bath was re-
moved, and the temperature of the reaction mixture increased
from 20 °C to 35–40 °C. The reaction mixture was stirred at this
temperature for 3 h and then maintained at 10 °C for 10 h. The so-
lid precipitated out was filtered on a Buchner funnel under suction
and washed with chilled methanol followed by diethyl ether to af-
ford potassium 4-phenyl-2-oxo-but-3-enoate as a yellow solid.
Acetyl chloride was added dropwisely to EtOH cooled in an ice bath
to produce dry hydrochloric acid. Potassium benzylidene pyruvate
was added at 0 °C. The reaction was warmed to room temperature
and stirred for 2 h and then refluxed for 6 h. Then extracted with
CH2Cl2, after the organic layer was separated and dried over NaSO4,
the solvent was evaporated, the crude product was recrystallized
in ethanol. Compounds 2p–s was prepared from the corresponding
aryl aldehydes and pyruvic acid.
4.1.2.1. 7,7-Dimethyl-2,4-diphenyl-4,6,7,8-tetrahydroquinolin-
5(1H)-one (3a). Yield (82.78%), mp 204–205 °C. ESIMS (m/
z) = 330.2 (M+H)+. IR (KBr) 3283.1, 3077.5, 2959.1, 1657.1, 1621.5,
1590.4, 1241.3, 762.6, 698.0 cmꢀ1 1H NMR (CDCl3, 300 MHz) d:
.
0.90 (s, 3H), 0.97 (s, 3H), 2.01–2.30 (m, 4H), 4.61 (d, J = 5.22 Hz,
1H), 5.16 (dd, J = 5.33 Hz, 1.7 Hz, 1H), 5.85 (s, 1H), 6.99–7.27 (m,
10H). 13C NMR (CDCl3, 75 MHz) d: 26.78, 28.89, 31.65, 37.27, 39.43,
50.35, 106.18, 106.42, 125.14, 125.32, 127.11, 127.52, 127.71,
127.82, 134.19, 135.37, 147.85, 152.14, 194.43. Anal. Calcd for
C23H23NO: C, 83.85; H, 7.04; N, 4.25. Found: C, 83.81; H, 7.01; N, 4.13.
4.1.2.2. 7,7-Dimethyl-4-(4-nitrophenyl)-2-phenyl-4,6,7,8-tetra-
hydroquinolin-5(1H)-one (3b). Yield (85.6%), mp 189–190 °C.
ESIMS (m/z) = 375.2 (M+H)+. IR (KBr); 3319.9, 3063.7, 1660.1,
1591.6, 838.4 cmꢀ1 1H NMR (CDCl3, 200 MHz) d: 0.99 (s, 3H),
.
1.10 (m, 3H), 2.24–2.31 (m, 2H), 2.39–2.49 (m, 2H), 4.60 (d,
J = 5.01 1H), 5.04 (dd, J = 5.01 Hz, J = 1.53, 1H), 6.76–6.82 (m, 2H),
7.15–7.33 (m, 7H), 7.53 (s, 1H). 13C NMR (CDCl3, 50 MHz) d:
26.37, 28.49, 31.21, 37.22, 39.65, 49.7, 103.74, 104.92, 122.37,
125.69, 126.46, 127.39, 127.55, 127.63, 128.07, 134.56, 135.03,
144.8, 152.13, 193.47. Anal. Calcd for C23H22N2O3: C, 73.78; H,
5.92; N, 7.48. Found: C, 73.82, H, 5.62, N, 7.39.
4.1.1.1. (E)-Ethyl 2-oxo-4-phenylbut-3-enoate (2o). Yield
(85.09%), ESIMS (m/z) = 205 (M+H)+. 1H NMR (CDCl3, 300 MHz) d:
1.40 (t, J = 7.2 Hz, 3H), 4.37 (q, J = 7.2 Hz, 2H), 7.36 (d, J = 16.0 Hz,
1H), 7.40–7.62 (m, 5H), 7.83 (d, J = 16.0 Hz, 1H). 13C NMR
(CDCl3,75 MHz,) d: 13.8, 62.2, 120.3, 128.77, 128.83, 131.4, 133.7,
148.1, 161.9, 182.6.
4.1.2.3. 4-(4-Chlorophenyl)-7,7-dimethyl-2-phenyl-4,6,7,8-tet-
rahydroquinolin-5(1H)-one (3c). Yield (85.6%), mp 207–209 °C.
ESIMS (m/z) = 375.2 (M+H)+. IR (KBr): 3286.7, 3076.1, 1654.8,

4144
A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
1567.0, 1210.3, 749.6 cmꢀ1
.
1H NMR (CDCl3, 200 MHz) d: 1.00 (s,
(m/z) = 320.1 (M+H)+. IR (KBr): 3319.9, 3063.7, 1660.1, 1591.6,
838.4 cmꢀ1 1H NMR (CDCl3 300 MHz) d: 1.79–1.86 (m, 2H),
3H), 1.10 (s, 3H), 2.00–2.15 (m, 2H), 2.52–2.56 (m, 2H) 4.61 (d,
J = 5.15 1H), 5.08 (dd, J = 5.15 Hz, J = 1.50, 1H), 7.19–7.97 (m, 9H),
8.32 (s, 1H). 13C NMR (CDCl3, 50 MHz) d: 28.18, 31.40, 35.13,
40.21, 43.77, 54.33, 109.75, 110.06, 118.09, 129.11, 131.97,
132.00, 132.61, 132.79, 136.54, 138.78, 147.45, 156.53, 198.31.
Anal. Calcd for C23H22ClNO: C, 75.92; H, 6.09; N, 3.85. Found: C,
75.99; H, 6.14; N, 3.81.
.
2.18–2.20 (m, 2H), 2.39–2.49 (m, 2H), 4.60 (d, J = 5.01 1H), 5.04
(dd, J = 5.01 Hz, J = 1.53, 1H), 6.76–6.82 (m, 2H), 7.15–7.33 (m,
7H), 7.53 (s, 1H). 13C NMR (CDCl3, 75 MHz) d: 21.12, 27.69, 36.82,
37.15, 106.40, 108.58, 125.75, 128.35, 128.43, 129.10, 129.21,
134.78, 135.82, 143.94, 143.98, 153.76, 162.63, 195.72. Anal. Calcd
for C21H18FNO: C, 78.98; H, 5.68; N, 4.39. Found: C, 79.06; H, 5.71;
N, 4.43.
4.1.2.4. 4-(4-Fluorophenyl)-7,7-dimethyl-2-phenyl-4,6,7,8-tet-
rahydroquinolin-5(1H)-one (3d). Yield (92.7%), ESIMS (m/
z) = 347.2 (M+H)+. IR (KBr); 3275.7, 3076.5, 1658.8, 1593.0,
4.1.2.10.
7,7-Dimethyl-2-phenyl-4-p-tolyl-4,6,7,8-tetrahydro-
quinolin-5(1H)-one (3j). Yield (79.4%), mp 206–208 °C. ESIMS
1222.3, 760.6 cmꢀ1 1H NMR (CDCl3 300 MHz) d: 0.99 (s, 3H),
.
(m/z) = 320.1 (M+H)+. IR (KBr): 3339.9, 3064.2, 1656.1, 1588.6,
1.08 (s, 3H), 2.03–2.48 (m, 4H), 4.63 (d, J = 5.22 Hz 1H), 5.15 (dd,
J = 5.22 Hz, J = 1.44 Hz 1H), 6.93–7.50 (m, 9H), 8.6 (s, 1H). 13C
NMR (CDCl3, 75 MHz) d: 30.81, 33.01, 35.73, 40.57, 44.26, 54.33,
109.81, 110.13, 118.10, 129.46, 132.01, 132.08, 132.73, 132.83,
138.57, 139.16, 148.15, 156.58, 198.36. Anal. Calcd for C23H22FNO:
C, 79.51; H, 6.38; N, 4.03. Found: C, 79.49, H, 6.42, N, 4.12.
838.4 cmꢀ1 1H NMR (CDCl3 300 MHz) d: 0.96 (s, 3H), 1.03 (s,
.
3H), 1.91- 2.20 (m, 4H), 2.30 (s, 3H), 4.55 (d, J = 5.28 Hz, 1H),
5.10 (dd, J = 5.28 Hz, 2H), 6.94–7.49 (m, 9H), 8.54 (s, 1H). 13C
NMR (CDCl3, 75 MHz) d: 21.18, 27.37, 29.68, 32.35, 37.68, 40.72,
50.89, 105.70, 106.48, 125.87, 126.00, 127.80, 128.48, 129.42,
132.81, 134.79, 138.14, 148.99, 152.88, 194.27. Anal. Calcd for
C24H25NO: C, 83.93; H, 7.34; N, 4.08: Found: C, 83.91; H, 7.48; N,
4.13.
4.1.2.5.
4-(4-Methoxyphenyl)-7,7-dimethyl-2-phenyl-4,6,7,8-
tetrahydroquinolin-5(1H)-one (3e). Yield (81.0%), mp 212–
214 °C. ESIMS (m/z) = 360.2 (M+H)+. IR (KBr): 3434.3, 3020.0,
4.1.2.11. 4-(1H-Indol-3-yl)-7,7-dimethyl-2-phenyl-4,6,7,8-tetra-
hydroquinolin-5(1H)-one (3k). Yield (79.4%), mp 240–242 °C.
ESIMS (m/z) = 369.2 (M+H)+. IR (KBr): 3381, 3284.1, 3076.4,
1605.5, 1216.2, 760.5, 670.0 cmꢀ1 1H NMR (CDCl3, 200 MHz) d:
.
0.92 (s, 3H), 1.00 (s, 3H), 2.08–2.26 (m, 4H), 3.66 (s, 3H), 4.58 (d,
J = 5.21 Hz, 1H), 5.18 (dd, J = 5.21 Hz, J = 1.56 Hz, 1H), 6.05 (s, 1H),
6.70–7.36 (m, 9H). 13C NMR (CDCl3, 50 MHz) d: 26.36, 28.38,
31.45, 35.81, 40.97, 49.76, 54.17, 106.21, 107.59, 112.64, 124.13,
127.50, 127.79, 132.93, 134.83, 139.21, 149.65, 156.83, 194.61.
Anal. Calcd for C24H25NO2: C, 80.19; H, 7.01; N, 3.90. Found: C,
80.13; H, 6.95; N, 3.91.
2956.2, 1655.6, 1623.2, 1591.3, 1241.3, 762.6, 698.1 cmꢀ1 1H
.
NMR (CDCl3, 200 MHz) d: 0.90 (s, 3H), 0.97 (S, 3H), 2.03–2.33 (m,
4H), 4.59 (d, J = 5.22 Hz, 1H), 5.14 (dd, J = 5.33 Hz, 1.7 Hz, 1H),
5.85 (s, 1H), 6.65–7.27 (m, 10H), 9.38 (s, 1H). 13C NMR (CDCl3,
50 MHz) d: 26.81, 28.89, 31.63, 37.26, 39.41, 50.34, 106.12,
106.41, 123.32, 125.14, 125.32, 127.11, 127.38, 127.52, 127.72,
127.83, 134.19, 135.38, 147.83, 152.11, 194.49. Anal. Calcd for
C25H24N2O: C, 81.49; H, 6.57; N, 7.60. Found: C, 81.53; H, 6.51;
N, 7.53.
4.1.2.6. 2-(4-Methoxyphenyl)-7,7-dimethyl-4-phenyl-4,6,7,8-tet-
rahydroquinolin-5(1H)-one (3f). Yield (78.7%), mp 214–216 °C.
ESIMS (m/z) = 360.0 (M+H)+. IR (KBr): 3274.7, 3076.5, 1668.8,
1573.0, 1221.3, 760.6 cmꢀ1
.
1H NMR (CDCl3, 200 MHz) d: 1.00 (s,
4.1.2.12. 2-Phenyl-4-p-tolyl-4,6,7,8-tetrahydroquinolin-5(1H)-
one (3l). Yield (77.9%), mp 203–205 °C. ESIMS (m/z) = 320.1
3H), 1.08 (s, 3H), 1.99–2.43 (m, 4H), 3.72 (s, 3H), 4.54 (d, J = 5.22 Hz,
1H), 5.10 (dd, J = 5.22 Hz, J = 1.28 Hz, 1H), 6.71 (d, J = 8.55 2H), 7.15
(d, J = 8.55, 2H), 7.29–7.48 (m, 5H), 8.35 (s, 1H). 13C NMR (CDCl3,
50 MHz) d: 26.11, 28.31, 30.94, 35.6, 37.5, 49.64, 53.75, 105.58,
105.8, 112.2, 124.68, 126.94, 127.09, 127.34, 133.36, 134.72,
139.72, 151.11, 156.33, 193.14. Anal. Calcd for C24H25NO2: C, 80.19;
H, 7.01; N, 3.90. Found: C, 80.18; H, 6.91; N, 3.98.
(M+H)+. IR (KBr): 3331.9, 3068.7, 1662.1, 1582.1, 843.5 cmꢀ1 1H
.
NMR (CDCl3, 300 MHz) d: 1.87–2.09 (m, 2H), 2.14–2.30 (m, 2H),
2.37 (s, 3H), 2.49–2.69 (m, 2H), 4.58 (d, J = 5.40 Hz, 1H), 5.18 (d,
J = 5.40, 1H), 7.07–7.46 (m, 9H), 8.61 (s, 1H). 13C NMR (CDCl3,
75 MHz) d: 21.17, 21.41, 27.37, 37.32, 37.38 105.60, 107.52,
125.89, 125.96, 127.74, 128.26, 128.53, 129.41, 132.79, 134.78,
138.16, 148.84, 154.77, 194.77. Anal. Calcd for C22H21NO: C,
83.78; H, 6.71; N, 4.44. Found: C, 83.85; H, 6.62; N, 4.48.
4.1.2.7. 2,4-Diphenyl-4,6,7,8-tetrahydroquinolin-5(1H)-one
(3g). Yield (81.5%), mp 206–209 °C. ESIMS (m/z) = 302.2 (M+H)+.
IR (KBr): 3247.3, 3063.5, 1653.3, 1586.5, 1488.9, 1272.2, 756.2,
4.1.2.13.
4-(4-Methoxyphenyl)-2-phenyl-4,6,7,8-tetrahydro-
693.0 cmꢀ1
.
1H NMR (CDCl3 200 MHz) d: 1.91–2.00 (m, 2H),
quinolin-5(1H)-one (3m). Yield (87.6%), mp 207–208 °C. ESIMS
2.25–2.32 (m, 2H), 2.57–2.67 (m, 2H), 4.66 (d, J = 5.32 1H), 5.14
(dd, J = 5.32 Hz, J = 1.57, 1H), 8.43 (s, 1H), 7.09–7.45 (m, 10H). Anal.
Calcd for C21H19NO: C, 83.69; H, 6.35; N, 4.65. Found: C, 83.78; H,
6.29; N, 4.61.
(m/z) = 332.3 (M+H)+. IR (KBr): 3275.7, 3074.5, 1658.8, 1593.0,
1222.3, 760.6 cmꢀ1 1H NMR (CDCl3, 200 MHz) d: 1.93–2.19 (m,
.
2H), 2.19–2.25 (m, 2H), 2.57–2.60 (m, 2H), 3.70 (s, 3H), 4.55 (d,
J = 5.34, 1H), 5.10 (dd, J = 5.34 Hz, J = 1.65, 1H), 7.13–7.84 (m,
9H), 8.50 (s, 1H). 13C NMR (CDCl3, 50 MHz) d: 19.78, 25.91, 35.06,
35.72, 53.53, 105.17, 106.72, 124.46, 126.70, 126.87, 127.10,
127.57, 128.11, 133.15, 134.43, 139.43, 156.12, 193.07. Anal. Calcd
for C22H21NO2: C, 79.73; H, 6.39; N, 4.23. Found: C, 79.61; H, 6.42;
N, 4.28.
4.1.2.8. 4-(4-Chlorophenyl)-2-phenyl-4,6,7,8-tetrahydroquino-
lin-5(1H)-one (3h). Yield (88.5%), mp 210 °C. ESIMS (m/
z) = 320.1 (M+H)+. IR (KBr): 3221.5, 1663.6, 1584.0, 757.6 cmꢀ1
.
1H NMR (CDCl3, 200 MHz) d: 0.85–1.24 (m, 2H), 2.22–2.28 (m,
2H), 2.54–2.60 (m, 2H), 4.63 (d, J = 5.26 1H), 5.07 (dd, J = 5.26 Hz,
J = 1.5 Hz, 1H), 7.14–8.05 (m, 9H), 8.46 (s, 1H). 13C NMR (CDCl3,
50 MHz) d: 20.16, 26.35, 36.04, 36.14, 104.74, 106.71, 124.95,
126.35, 127.40, 127.58, 128.04, 128.86, 129.83, 131.82, 134.9,
136.82, 141.79, 153.23, 193.75. Anal. Calcd for C21H18ClNO: C,
75.11; H, 5.40; N, 4.17. Found: C, 75.14; H, 5.34; N, 4.12.
4.1.2.14. 4-(Furan-2-yl)-7,7-dimethyl-2-phenyl-4,6,7,8-tetrahy-
droquinolin-5(1H)-one (3n). Yield (84.3%), mp 167–169 °C.
ESIMS (m/z) = 320 (M+H)+. IR (KBr): 3325.4, 3289.1, 3046.5,
2934.2, 1655.6, 1623.2 cmꢀ1 1H NMR (CDCl3, 200 MHz) d: 1.00
.
(s, 3H), 1.08 (s, 3H), 1.99–2.43 (m, 4H), 4.34 (d, J = 5.16 Hz, 1H),
5.06 (dd, J = 5.16 Hz, J = 1.28 Hz, 1H), 6.21 (d, J = 6.34, 1H), 7.29–
7.58 (m, 7H), 8.35 (s, 1H). Anal. Calcd for C21H21NO2: C, 78.97; H,
6.63; N, 4.39. Found: C, 78.49; H, 6.75; N, 4.48.
4.1.2.9. 4-(4-Fluorophenyl)-2-phenyl-4,6,7,8-tetrahydroquino-
lin-5(1H)-one (3i). Yield (74.5%), mp 202–204 °C. ESIMS

A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
4145
4.1.3. Typical experimental procedure for the synthesis of lactol
derivatives (3o–s)
4.1.4. Typical experimental procedure for the synthesis of ethyl
7,7-dimethyl-5-oxo-4-phenyl-1,4,5,6,7,8-hexahydroquinoline-
2-carboxylates derivatives (4o–s)
A solution of 3o (1 mmol) and ammonium acetate (2 mmol) in
MeOH (20 mL) was refluxed for 1 h. The solvent was evaporated
and solid was filtered and washed with cold ethanol to provide
4o as a yellow solid (97% yield).
Dimedone (1 mmol) and ethyl 4-phenyl-2-oxo-3-butenoate
(2o) at 0 °C under nitrogen atmosphere was added PTSA
20 mol % in THF. After 24 h the solvents were removed in vacuo.
The residue was purified by silica gel chromatography to afford
corresponding lactol derivatives as colorless oil of 3o in 78.5%
yield.
4.1.4.1. Ethyl 7,7-dimethyl-5-oxo-4-phenyl-1,4,5,6,7,8-hexahy-
4.1.3.1. Ethyl-2-hydroxy-7,7-dimethyl-5-oxo-4-phenyl-3,4,5,6,
7,8-hexahydro-2H-chromene-2-carboxylate (3o). Yield (78.5%),
ESIMS (m/z) = 345 (M+H)+. IR (KBr): 2965.2, 1745.3, 1625.6,
droquinoline-2-carboxylate (4o). Yield (97.0%), ESIMS (m/
z) = 356 (M+H)+. IR (KBr): 3331.7, 1686, 1662.3, 845.3 cmꢀ1 1H
.
NMR (CDCl3, 300 MHz) d: 1.02 (s, 3H), 1.08 (s, 3H), 1.31 (t,
J = 7.1 Hz, 3H), 2.19–2.35 (m, 4H), 4.25 (m, 2H), 4.72 (d,
J = 5.6 Hz, 1H), 6.18 (d, J = 5.6 Hz, 1H), 6.45 (s, 1H), 6.93–7.48 (m,
5H). 13C NMR (CDCl3, 75 MHz) d: 14.25, 27.64, 29.33, 32.68,
37.11, 41.81, 50.83, 55.33, 61.94, 107.54, 113.97, 117.47, 127.65,
132.23, 139.78, 152.01, 159.23, 165.9, 195.98. Anal. Calcd for
C20H23NO3: C, 73.82; H, 7.12; N, 4.30. Found: C, 73.91; H, 7.25;
N, 4.17.
1265.4, 1069.6 cmꢀ1 1H NMR (CDCl3, 300 MHz) d: 1.09 (s, 3H),
.
1.11 (s, 3H), 1.29 (t, J = 7.2 Hz, 3H), 2.21–2.61 (m, 6H), 3.89 (t,
J = 8.4 Hz, 1H), 4.17 (q, J = 7.2 Hz, 2H), 4.44 (s, 1H), 7.16–7.20 (m,
3H), 7.23–7.30 (m, 2H). 13C NMR (CDCl3, 75 MHz) d: 14.0, 27.83,
28.79, 31.76, 36.27, 38.31, 42.55, 50.86, 63.08, 95.04, 112.19,
126.14, 127.18, 143.20, 167.40, 167.91, 168.97, 196.55, 196.90.
Anal. Calcd for C20H24O5: C, 69.75; H, 7.02. Found: C, 69.59; H,
7.18.
4.1.4.2. Ethyl-4-(4-methoxyphenyl)-7,7-dimethyl-5-oxo-1,4,5,6,
4.1.3.2. Ethyl-2-hydroxy-4-(4-methoxyphenyl)-7,7-dimethyl-5-
oxo-3,4,5,6,7,8-hexahydro-2H-chromene-2-carboxylate
7,8-hexahydroquinoline-2-carboxylate
(4p). Yield
(95.2%),
ESIMS (m/z) = 356 (M+H)+. IR (KBr cmꢀ1); 3328.4, 1683, 1662.1,
1581.1, 845.7. 1H NMR (CDCl3, 300 MHz) d = 1.02 (s, 3H), 1.08 (s,
3H), 1.30 (t, J = 7.1 Hz, 3H), 2.18 (d, J = 16.4 Hz, 1H), 2.21 (d,
J = 16.4 Hz, 1H), 2.30 (d, J = 16.4 Hz, 1H), 2.35 (d, J = 16.4 Hz, 1H),
3.76 (s, 3H), 4.25 (m, 2H), 4.68 (d, J = 5.6 Hz, 1H), 6.11 (d,
J = 5.6 Hz, 1H), 6.44 (br s, 1H), 6.81 (d, J = 8.4 Hz, 2H), 7.20 (d,
J = 8.4 Hz, 2H), 13C NMR (CDCl3, 75 MHz): d = 14.24, 27.64, 29.33,
32.68, 37.06, 41.79, 50.83, 55.32, 61.93, 107.53, 113.95, 117.46,
125.77, 129.01, 138.78, 150.01, 158.33, 163.18, 195.50. Anal. Calcd
for C21H25NO4: C, 70.96; H, 7.09; N, 3.94. Found: C, 70.99; H, 7.17;
N, 4.08.
(3p). Yield (78.0%), ESIMS (m/z) = 375 (M+H)+. IR (KBr): 3456,
2960, 1745, 1623, 1070 cmꢀ1 1H NMR (CDCl3, 300 MHz) d: 1.08
.
(s, 3H), 1.11 (s, 3H), 1.27 (t, J = 7.2 Hz, 3H), 2.18–2.56 (m, 6H),
3.75 (s, 3H), 3.82–4.01 (m, 1H), 4.11–4.33 (m, 3H), 6.78 (d,
J = 6.82, 2H), 7.07 (d, J = 6.82, 2H). 13C NMR (CDCl3, 75 MHz) d:
14.03, 27.80, 29.42, 31.25, 36.04, 38.44, 42.54, 51.06, 55.26,
63.11, 95.89, 112.38, 114.33, 138.39, 135.06, 157.94, 166.94,
167.58, 169.11, 196.47, 196.92. Anal. Calcd for C21H26O6: C,
67.36; H, 7.00. Found: C, 67.43; H, 6.87.
4.1.3.3.
Ethyl-4-(4-chlorophenyl)-2-hydroxy-7,7-dimethyl-5-
oxo-3,4,5,6,7,8-hexahydro-2H-chromene-2-carboxylate
4.1.4.3. Ethyl 4-(4-chlorophenyl)-7,7-dimethyl-5-oxo-1,4,5,6,7,
8-hexahydroquinoline-2-carboxylate (4q). Yield (95.2%), ESIMS
(m/z) = 360 (M+H)+. IR (KBr): 3265.78, 1683.7, 1662.1,
(3q). Yield (79.4%), ESIMS (m/z) = 379 (M+H)+. IR (KBr): 3462,
2963, 1743, 1629, 1070 cmꢀ1 1H NMR (CDCl3, 300 MHz) d: 1.08
.
(s, 3H), 1.11 (s, 3H), 1.30 (t, J = 7.2 Hz, 3H), 2.16–2.67 (m, 6H),
3.82–4.01 (m, 1H), 4.19–4.42 (m, 3H), 7.12 (d, J = 7.13, 2H), 7.43
(d, J = 7.13, 2H). 13C NMR (CDCl3, 75 MHz) d: 14.09, 28.49, 29.43,
32.27, 38.13, 42.59, 51.13, 63.59, 94.88, 95.62, 129.14, 131.48,
143.69, 167.51, 168.2, 196.48, 197.95. Anal. Calcd for C20H23ClO5:
C, 63.41; H, 6.12: Found: C, 63.38; H, 6.19.
1583.2 cmꢀ1 1H NMR (CDCl3, 300 MHz) d:1.02 (s, 3H), 1.08 (s,
.
3H), 1.31 (t, J = 7.1 Hz, 3H), 2.23–2.41 (m, 4H), 4.26 (m, 2H), 4.74
(d, J = 5.6 Hz, 1H), 6.19 (d, J = 5.6 Hz, 1H), 6.51 (s, 1H), 7.19 (d,
J = 8.6 Hz, 2H), 7.39 (d, J = 8.6 Hz, 2H). Anal. Calcd for C20H22ClNO3:
C, 66.75; H, 6.16; N, 3.89. Found: C, 66.45; H, 6.84; N, 4.12.
4.1.4.4. Ethyl-4-(4-fluorophenyl)-7,7-dimethyl-5-oxo-1,4,5,6,7,
8-hexahydroquinoline-2-carboxylate (4r). Yield (87.8%), ESIMS
(m/z) = 344 (M+H)+. IR (KBr): 3275.7, 3076.2, 1661.4,
4.1.3.4.
Ethyl-4-(4-fluorophenyl)-2-hydroxy-7,7-dimethyl-5-
oxo-3,4,5,6,7,8-hexahydro-2H-chromene-2-carboxylate
(3r). Yield (84.8%), ESIMS (m/z) = 363 (M+H)+. IR (KBr): 3468,
1594.3 cmꢀ1 1H NMR (CDCl3, 300 MHz) d: 1.02 (s, 3H), 1.08 (s,
.
2969, 1745, 1637, 1076 cmꢀ1
.
1H NMR (CDCl3, 300 MHz) d: 1.08
3H), 1.31 (t, J = 7.1 Hz, 3H), 2.13–2.48 (m, 4H), 4.28 (m, 2H), 4.79
(d, J = 5.6 Hz, 1H), 6.23 (d, J = 5.6 Hz, 1H), 6.53 (s, 1H), 7.21 (d,
J = 8.6 Hz, 2H), 7.42 (d, J = 8.6 Hz, 2H). Anal. Calcd for C20H22FNO3:
C, 69.95; H, 6.46; N, 4.08. Found: C, 70.12; H, 6.58; N, 3.97.
(s, 3H), 1.11 (s, 3H), 1.30 (t, J = 7.2 Hz, 3H), 2.19–2.69 (m, 6H),
3.86–4.09 (m, 1H), 4.19–4.42 (m, 3H), 7.17–7.59 (m, 4H). 13C
NMR (CDCl3, 75 MHz) d: 14.09, 28.50, 29.43, 32.29, 38.18, 42.61,
51.15, 63.59, 94.88, 95.62, 120.21, 131.79, 143.84, 167.62, 168.37,
196.53, 197.99. Anal. Calcd for C20H23FO5: C, 66.29; H, 6.40. Found:
C, 66.38; H, 6.29.
4.1.4.5. Ethyl-4-(4-bromophenyl)-7,7-dimethyl-5-oxo-1,4,5,6,7,
8-hexahydroquinoline-2-carboxylate (4s). Yield (92.8%), ESIMS
(m/z) = 404 (M+H)+. IR (KBr): 3254.3, 3073.5, 1661.6 cmꢀ1 1H
.
4.1.3.5.
Ethyl-4-(4-bromophenyl)-2-hydroxy-7,7-dimethyl-5-
NMR (CDCl3, 300 MHz) d: 1.02 (s, 3H), 1.08 (s, 3H), 1.31 (t,
J = 7.1 Hz, 3H), 2.12–2.43 (m, 4H), 4.28 (m, 2H), 4.75 (d,
J = 5.6 Hz, 1H), 6.20 (d, J = 5.6 Hz, 1H), 6.53 (s, 1H), 7.16 (d,
J = 8.8 Hz, 2H), 7.67 (d, J = 8.8 Hz, 2H). Anal. Calcd for C20H22BrNO3:
C, 59.42; H, 5.48; N, 3.46. Found: C, 59.19; H, 5.49; N, 3.32.
oxo-3,4,5,6,7,8-hexahydro-2H-chromene-2-carboxylate
(3s). Yield (92.1%), ESIMS (m/z) = 423 (M+H)+. IR (KBr). 3440,
3053, 1745, 1070 cmꢀ1 1H NMR (CDCl3, 300 MHz) d: 1.08 (s, 3H),
.
1.11 (s, 3H), 1.29 (t, J = 7.2 Hz, 3H), 2.14–2.62 (m, 6H), 3.82–4.01
(m, 1H), 4.17–4.39 (m, 3H), 7.03 (d, J = 6.95, 2H), 7.34 (d, J = 6.95,
2H). 13C NMR (CDCl3, 75 MHz) d: 14.08, 27.81, 29.44, 31.72,
33.23, 35.54, 38.13, 42,56, 50.96, 63.39, 94.84, 111.84, 113.95,
119.88, 128.95, 131.30, 142.48, 143.56, 167.34, 167.96, 169.00,
196.35, 196.81. Anal. Calcd for C20H23BrO5: C, 56.75; H, 5.48.
Found: C, 56.70; H, 5.86.
4.1.5. Typical experimental procedure for the synthesis of 7,7-
dimethyl-5-oxo-4-phenyl-1,4,5,6,7,8-hexahydroquinoline-
2-carboxylic acid derivatives (5a–e)
Compound 4o (1 mmol) was dissolved in EtOH (10 mL and ta-
ken in to the 50 mL R.B. flask and cool to 0 °C. 20% sodium carbon-

4146
A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
ate solution of EtOH was added dropwise to compound 4o and stir-
red for 20 h. After the solid appeared was filtered and washed with
cold ethanol to give 5a in 96.7% yield.
4.1.6.1. 4-(4-Fluorophenyl)-7,7-dimethyl-2-phenyl-7,8-dihy-
droquinolin-5(6H)-one (6a). Yield (82.3%), mp 176–178 °C.
ESIMS (m/z) = 346 (M+H)+. IR (KBr): 3074.6, 1656.2, 1591.5 cmꢀ1
.
1H NMR (CDCl3, 200 MHz) d: 1.01 (s, 3H), 1.08 (s, 3H), 2.04- 2.43
(m, 4H), 7.08–7.56 (m, 7H), 7.62–7.96 (m, 3H). 13C NMR (CDCl3,
50 MHz) d: 24.23, 24.37, 32.55, 43.35, 49.48, 112.32, 113.46,
118.84, 124.10, 124.21, 126.18, 127.35, 132.31, 133.21, 138.29,
144.26, 146.33, 152.17, 159.47, 198.29. Anal. Calcd for C23H20FNO:
C, 79.98; H, 5.84; N, 4.06. Found: C, 79.74; H, 5.69; N, 4.22.
4.1.5.1. 7,7-Dimethyl-5-oxo-4-phenyl-1,4,5,6,7,8-hexahydro-
quinoline-2-carboxylic acid (5a). Yield (96.7%), ESIMS (m/
z) = 298 (M+H)+. IR (KBr): 3456–3113.7, 1686, 1659.3,
845.3 cmꢀ1 1H NMR (CD3OD, 300 MHz) d: 1.00 (s, 3H), 1.06 (s,
.
3H), 2.19–2.35 (m, 4H), 4.69 (d, J = 5.6 Hz, 1H), 6.17 (d, J = 5.6 Hz,
1H), 6.45 (s, 1H), 6.89–7.42 (m, 5H), 12.34 (s, 1H). 13C NMR
(CD3OD, 75 MHz) d: 27.64, 29.33, 32.65, 37.11, 41.81, 50.83,
55.33, 107.51, 113.96, 117.47, 127.66, 130.21, 138.56, 152.01,
159.23, 161.67, 194.78. Anal. Calcd for C18H19NO3: C, 72.71; H,
6.44; N, 4.71. Found: C, 72.71; H, 6.44; N, 4.71.
4.1.6.2. 4-(4-Chlorophenyl)-7,7-dimethyl-2-phenyl-7,8-dihy-
droquinolin-5(6H)-one (6b). Yield (82.3%), mp 188–189 °C.
ESIMS (m/z) = 362 (M+H)+. IR (KBr): 3074.2, 1655.4, 1588.2, 956,
742 cmꢀ1 1H NMR (CDCl3, 200 MHz) d: 1.01 (s, 3H), 1.08 (s, 3H),
.
2.03–2.43 (m, 4H), 6.93–7.42 (m, 7H), 7.39–7.61 (m, 2H), 7.79 (s,
1H). 13C NMR (CDCl3, 50 MHz) d: 24.23, 24.35, 32.55, 43.31,
49.48, 114.24, 115.16, 118.81, 124.23, 125.67, 128.18, 132.31,
137.24, 138.37, 144.73, 147.33, 153.21, 159.47, 198.29. Anal. Calcd
for C23H20ClNO: C, 76.34; H, 5.57; N, 3.87. Found: C, 76.42; H, 5.64;
N, 4.01.
4.1.5.2. 4-(4-Methoxyphenyl)-7,7-dimethyl-5-oxo-1,4,5,6,7,8-
hexahydroquinoline-2-carboxylic acid (5b). Yield (92.8%),
ESIMS (m/z) = 328 (M+H)+. IR (KBr): 3412–3132.4, 1684.5, 1656.1,
1576.9 cmꢀ1 1H NMR (CD3OD, 300 MHz) d: 1.00 (s, 3H), 1.06 (s,
.
3H), 2.16 (d, J = 16.4 Hz, 1H), 2.21 (d, J = 16.4 Hz, 1H), 2.30 (d,
J = 16.4 Hz, 1H), 2.35 (d, J = 16.4 Hz, 1H), 3.89 (s, 3H), 4.68 (d,
J = 5.6 Hz, 1H), 6.11 (d, J = 5.6 Hz, 1H), 6.79 (d, J = 8.1 Hz, 2H),
7.16 (d, J = 8.1 Hz, 2H), 10.24 (s, 1H), 12.89 (s, 1H). 13C NMR
(CD3OD, 75 MHz) d: 27.63, 29.33, 32.65, 37.06, 41.77, 50.78,
55.65, 107.57, 113.75, 117.18, 125.77, 128.88, 138.78, 150.01,
158.32, 159.39, 195.24. Anal. Calcd for C19H21NO4: C, 69.71; H,
6.47; N, 4.28. Found: C, 69.62; H, 6.38; N, 4.11.
4.1.6.3. 4-(4-Methoxyphenyl)-7,7-dimethyl-2-phenyl-7,8-dihy-
droquinolin-5(6H)-one (6c). Yield (82.3%), mp 220–222 °C.
ESIMS (m/z) = 358 (M+H)+. IR (KBr): 3070.3, 1654.2, 1588.5 cmꢀ1
.
1H NMR (CDCl3, 200 MHz) d: 1.01 (s, 3H), 1.08 (s, 3H), 2.03–2.39
(m, 4H), 3.66 (s, 3H), 6.57–7.47 (m, 7H), 7.39–7.58 (m, 2H), 7.77
(s, 1H). 13C NMR (CDCl3, 50 MHz) d: 24.22, 24.33, 32.55, 43.32,
49.48, 54.12, 111.23, 114.78, 118.65, 124.13, 125.62, 128.11,
131.89, 137.23, 138.33, 142.39, 144.73, 147.26, 153.21, 159.47,
197.86. Anal. Calcd for C24H23NO2: C, 80.64; H, 6.49; N, 3.92.
Found: C, 76.42; H, 5.64; N, 4.01.
4.1.5.3. 4-(4-Chlorophenyl)-7,7-dimethyl-5-oxo-1,4,5,6,7,8-hex-
ahydroquinoline-2-carboxylic acid (5c). Yield (96.2%), ESIMS
(m/z) = 332 (M+H)+. IR (KBr cmꢀ1); 3417.8–3119.6, 1683.7,
1656.2, 1585.9. 1H NMR (CD3OD, 300 MHz) d: 1.00 (s, 3H), 1.06
(s, 3H), 2.19–2.36 (m, 4H), 4.73 (d, J = 5.45 Hz, 1H), 6.13 (d,
J = 5.45 Hz, 1H), 7.14 (d, J = 8.39 Hz, 2H), 7.33 (d, J = 8.39 Hz, 2H).
10.45 (s, 1H), 12.68 (s, 1H). Anal. Calcd for C18H18ClNO3: C,
65.16; H, 5.47; N, 4.22. Found: C, 65.04; H, 5.12; N, 4.06.
4.2. Pharmacology
4.2.1. Sucrose loaded rat model (SLM)
Male albino rats of Charles Foster/Wistar strain of average body
weight 160 20 g were selected for this study. The blood glucose
level of each animal was checked by glucometer using glucostrips
(Boehringer Mannheim) after 16 h starvation. Animals showing
blood glucose levels between 3.33 and 4.44 mM (60–80 mg/dl)
were divided into groups of five to six animals in each. Animals
of experimental group were administered suspension of the de-
sired synthetic compound orally (made in 1.0% gum acacia) at a
dose of 100 mg/kg body weight. Animals of control group were gi-
ven an equal amount of 1.0% gum acacia. A sucrose load (10.0 g/kg)
was given to each animal orally exactly after 30 min post adminis-
tration of the test sample/vehicle. Blood glucose profile of each rat
was again determined at 30, 60, 90, and 120 min post administra-
tion of sucrose by glucometer Food but not water was withheld
from the cages during the course of experimentation. Quantitative
glucose tolerance of each animal was calculated by Area Under
Curve (AUC) method (Prism Software). Comparing the AUC of
experimental and control groups determined the percentage anti-
hyperglycemic activity.
4.1.5.4. 4-(4-Fluorophenyl)-7,7-dimethyl-5-oxo-1,4,5,6,7,8-hexa-
hydroquinoline-2-carboxylic acid (5d). Yield (79.8%), ESIMS (m/
z) = 316 (M+H)+. IR (KBr): 3424.6–3123.9, 3278.5, 3079.3,
1654.9 cmꢀ1 1H NMR (CD3OD, 300 MHz) d: 1.00 (s, 3H), 1.06 (s,
.
3H), 2.13–2.48 (m, 4H), 4.74 (d, J = 5.58 Hz, 1H), 6.21 (d, J = 5.58 Hz,
1H), 7.20 (d, J = 8.59 Hz, 2H), 7.41 (d, J = 8.59 Hz, 2H), 10.38 (s, 1H),
12.56 (s, 1H). 13C NMR (CDCl3, 75 MHz) d: 27.69, 29.34, 32.69, 36.99,
41.84, 50.83, 113.98, 117.46, 125.77, 128.76, 138.69, 149.82, 158.31,
156.11, 195.17. Anal. Calcd for C18H18FNO3: C, 68.56; H, 5.75; N,
4.44. Found: C, 68.49; H, 5.83; N, 4.11.
4.1.5.5. 4-(4-Bromophenyl)-7,7-dimethyl-5-oxo-1,4,5,6,7,8-hex-
ahydroquinoline-2-carboxylic acid (5e). Yield (82.3%), ESIMS
(m/z) = 404 (M+H)+. IR (KBr cmꢀ1); 3425.7–3124.3, 3279.3,
3081.9, 1658.1, 1555.8. 1H NMR (CD3OD, 300 MHz) d: 1.00 (s,
3H), 1.06 (s, 3H), 2.15–2.41 (m, 4H), 4.74 (d, J = 5.57 Hz, 1H), 6.19
(d, J = 5.57 Hz, 1H), 7.19 (d, J = 8.74 Hz, 2H), 7.69 (d, J = 8.74 Hz,
2H), 10.29 (s, 1H), 12.63 (s,1H). Anal. Calcd for C18H18BrNO3: C,
57.46; H, 4.82; N, 3.72. Found: C, 57.49; H, 4.75; N, 3.81.
4.2.2. Sucrose-challenged streptozotocin-induced diabetic rat
model (STZ-S)
4.1.6. Typical experimental procedure for the aromatization of
polyhydroquinoline (6a–c)
Male albino rats of Sprague Dawley strain of body weight
160 20 g were selected for this study. Streptozotocin (Sigma,
USA) was dissolved in 100 mM citrate buffer pH 4.5 and calculated
amount of the fresh solution was injected to overnight fasted rats
(60 mg/kg) intraperitoneally. Blood glucose level was checked 48 h
later by glucostrips and animals showing blood glucose values be-
tween 144 and 270 mg/dl (8–15 mM) were included in the exper-
iment and considered as diabetic. The diabetic animals were
divided into groups consisting of five to six animals in each group.
Polyhydroquinoline 3d (1 mmol), HbA (0.1 mmol), H2O2 (30%
w/v, 2 mmol), 15% acetonitrile in phosphate buffer (pH 6.5, 3 mL
was taken in to the 50 mL R.B. flask and stirred at room tempera-
ture for 12 h. After the completion of the reaction acetonitrile
was evaporated and diluted with DCM (125 mL) and washed with
water (3 ꢁ 50 mL). Obtained crude product was purified by silica
gel column chromatography in 82% yield.

A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
4147
Animals of experimental groups were administered suspension of
the desired test samples orally (made in 1.0% gum acacia) at a dose
of 100 mg/kg body weight. Animals of control group were given an
equal amount of 1.0% gum acacia. A sucrose load of 2.5 g/kg body
weight was given after 30 min of compound administration. After
30 min of post sucrose load blood glucose level was again checked
by glucostrips at 30, 60, 90, 120, 180, 240, 300 min, and at 24 h,
respectively. Animals not found diabetic after 24 h post treatment
of the test sample were not considered and omitted from the cal-
culations and termed as non-responders. The animals, which did
not show any fall in blood glucose profile in a group while the oth-
ers in that group, showed fall in blood glucose profile were also
considered as non-responders. Food but not water was withheld
from the cages during the experimentation. Comparing the AUC
of experimental and control groups determined the percent anti-
hyperglycemic activity.
4.2.4. Protein-tyrosine phosphatase 1B inhibitory assay
The test compounds were pre-incubated for 10 min with the
enzyme in the absence and presence of 0.01% Triton X–100. Assay
was performed in final volume of 1.0 mL in test mixture containing
10 mM of pNPP in 50 mM HEPES buffer (pH 7.0) with 1 mM DTT
and 2 mM EDTA. The reaction was stopped after 30 min of incuba-
tion at 37 °C by addition of 500
ll of 0.1 N NaOH and the absor-
bance was determined at 410 nm. A molar extinction coefficient
of 1.78 ꢁ 104 Mꢀ1 cmꢀ1 was utilized to calculate the concentration
of p-nitrophenolate ion generated in the reaction mixture. Sodium
orthovanadate was taken as standard in enzyme assay. The IC50 of
the compounds were determined by constructing a dose–response
curve and examining the effect of different concentrations of
compounds.
4.2.5. Glucose-6-phosphatase inhibitory activity assay
The 1.0 mL assay system contained 0.3 M citrate buffer (pH 6.0),
28 mM EDTA, 14 mM NaF, 200 mM glucose-6-phosphate, and en-
zyme protein. The mixture was incubated at 37 °C for 30 min after
which 1.0 mL of 10% TCA was added. Glucose-6-phosphatase activ-
ity was defined as micromole Pi released per minute per milligram
protein.
4.2.3. Antihyperglycemic activity in db/db mice
The background for the db/db mouse is the C57BL/Ks strain. The
major deficiency of the C57BL/KsBom-db mouse (db/db) is lack of a
functional leptin receptor. This leads to defective leptin signaling
and a complete lack of feedback from leptin. Both hypothalamic
Neuropeptide Y (NPY) content and secretion are consequently
elevated, and this result in hyperphagia and decreased energy
expenditure, obesity, insulin-resistance, hyperinsulinemia, hyper-
glycaemia and dyslipidemia. The db/db mouse develops non insu-
lin dependent diabetes mellitus (NIDDM) from around week 10.
The disease is stable until week 20, where destruction of pancreatic
b-cells can be recognized clinically as decreasing levels of plasma
insulin and very severe hyperglycaemia. The db/db mouse has a
maximal life span of 9–12 months. The male mice are more dia-
betic than, and will normally die earlier than the females. The
advantage of using male mice for experimental purposes is that
the fluctuations in plasma parameters are less than in the females
where the oestrogen cycle affects the clinical diabetes. The optimal
age of db/db mice used for experiments is from week 12 to 18
when they have developed non insulin dependent diabetes melli-
tus (NIDDM) with diabetic dyslipidemia but still have functional
b-cells in the pancreas. C57BL/KsBom-db mice 12–18 weeks, 40–
50 g bred in the animal house of CDRI, Lucknow. Ten mice (5 males
and 5 females) were used in the experiments. The mice were
housed in groups of 5 (same sex) in a room controlled for temper-
ature (23 2.0 °C) and 12/12 h light/dark cycle (lights on at
6.00 am). Body weight was measured daily from day 1 to day 10.
All animals had free access to fresh water and to normal chow ex-
cept on the days of the postprandial protocol day 6 and during the
overnight fast before the OGTT on day 10. The animals always had
access to water during experimental periods. Blood glucose was
checked every morning up till day 5. On day 6 postprandial proto-
col was employed, in this method blood glucose was checked at
ꢀ0.5 h and 0 h. Test drugs were given to the treatment group
whereas vehicle received only gum acacia (1.0%); the blood glucose
was again checked at 1, 2, 3, 4, and 6 h post test drug treatment. On
day 10 an oral glucose tolerance test (OGTT) was performed after
an overnight fasting. Blood glucose was measured at ꢀ30.0 min
and test drugs were administered. The blood glucose was again
measured at 0.0 min post treatment and at this juncture glucose
solution was given at a dose of 3 gm/kg to all the groups including
vehicle. The blood glucose levels were checked at 30 min, 60 min,
90 min, and 120 min post glucose administration. At the end of
the experiment blood has been withdrawn from the retro-orbital
plexus of mice for the estimation of serum insulin. Quantitative
glucose tolerance of each animal was calculated by area under
curve (AUC) method (Prism software). Comparing the AUC of
experimental and control groups determined the %antihyperglyce-
mic activity. Statistical comparison was made by Dunnett’s test.
4.2.6. Glycogen phosphorylase inhibitory activity assay
Mixture A contained 57 mg glycogen (substrate), 188 mg glu-
cose-1-phosphate, 42 mg sodium fluoride, 138.8 mg 50-AMP
(4 mM) dissolved in 10 mL water. The reaction mixture containing
0.2 mL Mixture-A, 0.1 mL enzyme is incubated for 30 min at 37 °C
and reaction terminated by adding 0.1 mL TCA (10%) and 0.4 mL of
0.1 M sodium acetate. The mixture is kept overnight at 4 °C and the
Pi released is estimated.
4.2.7.
a-Glucosidase inhibitory activity assay
a-Glucosidase inhibitory activity was determined in 1 mL reac-
tion system in 67 mM sodium phosphate buffer (pH 6.8) contain-
ing 1.0 mg/mL glutathione in the presence of 0.1 mg/mL purified
a-glucosidase. The reaction was started by adding 3.0 mg/mL
pNPG to the reaction mixture. The reaction was followed for
3 min at 405 nm at the interval of 30 s.
4.2.8. DPP IV enzyme inhibition assay
Enzymatic activity was determined at 37 °C by the cleavage rate
of a substrate, Gly-Pro-AMC (30 lM) (Sigma–Aldrich, USA). Briefly,
10
bottomed microtiter plate, followed by the addition of 50
60 L Gly-Pro-AMC, 10 L of 500 mM Tris–HCl (pH 7.4), 20
distilled water, and 10 L of a test compound. The change of fluo-
l
L of DPP-IV solution was added to each well of a 96-well flat-
L of
L of
l
l
l
l
l
rescence was monitored at 37 °C using a spectrofluorometer (exci-
tation at 355 nm/emission at 460 nm) (fmax, Molecular Devices,
USA). The initial rate of DPP-IV enzyme activity was calculated
over the first 15 min of the reaction, with units/mL being defined
as the rate of increase in the fluorescence intensity (arbitrary units)
under these conditions.
4.2.9. Antidyslipidemic activity
Adult male Charles Foster rats (200 225 g) bred in the animal
house of the institute were used for the lipid lowering activity. Rats
were divided in control, triton induced, triton plus compounds and
Gemfibrozil (100 mg/kg) treated groups containing five rats in
each. Hyperlipidemia was developed by administration of Triton
WR-1339 (Sigma chemical co., St. Louis, USA) at a dose of
400 mg/kg body wt intraperitoneally to animals of all groups ex-
cept the control. Compounds were macerated with gum acacia
(0.2% w/v), suspended in water and fed simultaneously with triton
at a dose of 100 mg/kg po to the animals of treated groups. Animals
of the control and triton group without treatment with test com-

4148
A. Kumar et al. / Bioorg. Med. Chem. 18 (2010) 4138–4148
12. Hilgeroth, A. Mini-Rev. Med. Chem. 2002, 2, 235.
13. Kawase, M.; Shah, A.; Gaveriya, H.; Motohashi, N.; Sakagami, H.; Varga, A.;
Molnar J. Bioorg. Med. Chem. 2002, 10, 1051.
pounds were given same amount of gum acacia suspension (vehi-
cle). After 18 h of treatment (50 mg/kg b. wt) 1.0 mL blood was
withdrawn from retro-orbital sinus using glass capillary in EDTA
coated eppendorf tube (3.0 mg/mL blood). The blood was centri-
fuged (at 2500g) at 4 °C for 10 min and the plasma was separated.
Plasma was diluted with normal saline (ratio 1:3) and used for
analysis of total cholesterol (TC), phospholipids (PL), and triglycer-
ides (TG) by standard enzymatic procedures.
14. (a) Kumar, A.; Maurya, R. A.; Sharma, S.; Ahmad, P.; Singh, A. B.; Tamrakar, A.
K.; Srivastava, A. K. Bioorg. Med. Chem. 2009, 17, 5285–5292; (b) Kumar, A.;
Ahmad, P.; Maurya, R. A.; Singh, A. B.; Tamrakar, A. K.; Srivastava, A. K. Eur. J.
Med. Chem. 2009, 44, 109; Kumar, A.; Maurya, R. A.; Sharma, S.; Ahmad, P.;
Singh, A. B.; Bhatia, G.; Srivastava, A. K. Bioorg. Med. Chem. Lett. 2009.
doi:10.1016/j.bmcl.2009.09.031.
15. (a) Ladouceur, G. H.; Cook, J. H.; Doherty, E. M.; Schoen, W. R.; MacDougall, M.
L.; Livingston, J. N. Bioorg. Med. Chem. Lett. 2002, 12, 461; (b) Schoen, W. R.;
Ladouceur, G. H.; Cook, J. H.; Lease, T. G.; Wolanin, D. J.; Kramss, R. H.; Hertzog,
D. L.; Osterhout, M. H. U.S. Patent No. 6218,431, 2001.
Acknowledgements
16. (a) Goldmann, S.; Ahr, H.-J.; Puls, W.; Bischoff, H.; Petzinna, D.; Schlossman, K.;
Bender, J. U.S. Patent, 4786,641, 1988; (b) Oikonomakos, N. G.; Tsitsanou, K. E.;
Zographos, S. E.; Skamnaki, V. T.; Goldmann, S.; Bischoff, H. Protein Sci. 1999, 8,
1930; (c) Bergans, N.; Stalmans, W.; Goldmann, S.; Vanstapel, F. Diabetes 2000,
49, 1419.
S.S, V.D.T, R.A.M, and S.P.S are thankful to CSIR New Delhi for
the award of fellowship. The authors also acknowledge CSIR net-
working project NWP-032 for financial support and highly thankful
to SAIF for providing the spectral data.
17. Poindexter Swann, G. S.; Swann, R. T.; Bruce, M. A.; Morton, M. A.; Huang, Y.;
Sit, S.; Breitenbucher, J. G. WO/1998/054136, 1998.
18. Sogawa, S.; Nihro, Y.; Ueda, H.; Izumi, A.; Miki, T.; Matsumoto, H.; Satoh, T. J.
Med. Chem. 1993, 36, 3904.
19. Kumar, A.; Maurya, R. A.; Sharma, S. Bioorg. Med. Chem. Lett. 2009, 19, 4432.
20. Taussky, H. H.; Shorr, E. A. J. Biol. Chem. 1953, 202, 675.
21. Hubscher, G.; West, G. R. Nature 1965, 205, 799.
22. Wright, S. W.; Ammirati, M. J.; Andrews, K. M.; Brodeur, A. M.; Danley, D. E.;
Doran, S. D.; Lillquist, J. S.; McClure, L. D.; McPherson, R. K.; Orena, S. J.; Parker,
J. C.; Polivkova, J.; Qiu, X.; Soeller, W. C.; Soglia, C. B.; Treadway, J. L.;
VanVolkenberg, M. A.; Wang, H.; Wilder, D. C.; Olson, T. V. J. Med. Chem. 2006,
49, 3068.
23. Rall, T. W.; Sutherland, E. W.; Barthet, J. J. Biol. Chem. 1957, 244, 463.
24. Lebovitz, H. E. Endocrinol. Metab. Clin. North Am. 1997, 26, 539–551.
25. Goldstein, B. J.; Bittner-Kowalezyk, A.; White, M. F.; Harbeck, M. J. Biol. Chem.
2000, 275, 4283.
References and notes
1. (a) Tassoni, E.; Giannessi, F.; Brunetti, T.; Pessotto, P.; Renzulli, M.; Travagli, M.;
Rajamäki, S.; Prati, S.; Dottori, S.; Corelli, F.; Cabri, W.; Carminati, P.; Botta, M. J.
Med. Chem. 2008, 51, 3073; (b) Ross, S. A.; Gulve, E. A.; Wang, M. Chem. Rev.
2004, 104, 1255; (c) Rakowitz, D.; Maccri, R.; Ottana, R.; Vigorita, M. G. Bioorg.
Med. Chem. Lett. 2006, 14, 567.
2. King, H.; Aubert, R. E.; Herman, W. H. Diabetes Care 1998, 21, 1414.
3. Boyle, J. P.; Honeycutt, A. A.; Narayan, V. K. M.; Hoerger, T. J.; Geiss, L. S.; Chen,
H.; Thompson, T. J. Diabetes Care 2001, 24, 1936.
4. Tackling the threat to health of diabetes mellitus (Editorial). Lancet 2006, 398,
1624.
5. Wild, S.; Roglic, G.; Green, A.; Sicree, R.; King, H. Diabetes Care 2004, 27, 1047.
6. Zimmet, P. Z.; Alberti, K. G. M. M.; Shaw, J. Nature 2001, 414, 782.
7. American Diabetes Association Diabetes Care 2003, 26, 917–932.
8. Pyorala, K.; Pedersen, T. R.; Kjekshus, J.; Fraergeman, O.; Olsson, A. G.;
Thorgeirsson, G. Diabetes Care 1997, 20, 614.
27. Kumar, A.; Maurya, R. A. Tetrahedron 2007, 64, 3477.
Further reading
9. (a) Bossert, F.; Meyer, H.; Wehinger, E. Angew. Chem., Int. Ed. Engl. 1981, 20, 762;
(b) Nakayama, H.; Kasoaka, Y. Heterocycles 1996, 42, 901.
10. Triggle, D. J.; Langs, D. D.; Janis, R. A. Med. Res. Rev. 1989, 9, 123.
11. (a) Mason, R. P.; Mak, I. T.; Trumbore, M. W.; Mason, P. E. Am. J. Cardiol. 1999,
84, 16–22; (b) Aruoma, O.; Smith, C.; Cecchini, R.; Evans, P.; Halliwell, B.
Biochem. Pharmacol. 1991, 42.
26. (a) Schurr, P. E.; Schultz, J. R.; Parkinson, T. M. Lipids 1972, 7, 68; (b) Deeg, R.;
Ziegehorn J. Clin. Chem. 1983, 29, 1798.