Pyranocoumarins: A new class of anti-hyperglycemic and anti-dyslipidemic agents

Article abstract of DOI:10.1016/j.bmcl.2009.09.031

A series of pyranocoumarin derivatives were synthesized and evaluated in vivo for their anti-hyperglycemic as well as anti-dyslipidemic activities. Compounds 7a, 7c, 8a, 8b, 8c, 8e and 8f have shown promising anti-hyperglycemic activities in sucrose loaded model (SLM) as well as sucrose challenged streptozotocin induced diabetic rat model (STZ). Compounds 8a and 8b were showing 38.0% and 42.0% blood glucose lowering activity in db/db mice model. In vitro anti-hyperglycemic activity evaluation exhibited that compounds 8a (IC₅₀ = 24.5 μM) and 8b (IC₅₀ = 36.2 μM) are potential PTP-1B inhibitors thereby revealing their possible mechanism of anti-diabetic action. Compounds 7a, 7b, 8a, 8b, 8d, 8e and 8f have shown significant anti-dyslipidemic activity in triton induced dyslipidemia in rats.

Full text of DOI:10.1016/j.bmcl.2009.09.031

Bioorganic & Medicinal Chemistry Letters 19 (2009) 6447–6451
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Pyranocoumarins: A new class of anti-hyperglycemic
and anti-dyslipidemic agents
a
a
a
b
b
Atul Kumar a, , Ram Awatar Maurya , Siddharth Sharma , Pervez Ahmad , A. B. Singh , Gitika Bhatia ,
*
Arvind K. Srivastava b
a Medicinal and Process Chemistry Division, Central Drug Research Institute, Council of Scientific & Industrial Research, Lucknow 226 001, India
b Biochemistry Division, Central Drug Research Institute, Council of Scientific & 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:
Received 23 April 2009
Revised 7 September 2009
Accepted 9 September 2009
Available online 12 September 2009
A series of pyranocoumarin derivatives were synthesized and evaluated in vivo for their anti-hypergly-
cemic as well as anti-dyslipidemic activities. Compounds 7a, 7c, 8a, 8b, 8c, 8e and 8f have shown prom-
ising anti-hyperglycemic activities in sucrose loaded model (SLM) as well as sucrose challenged
streptozotocin induced diabetic rat model (STZ). Compounds 8a and 8b were showing 38.0% and 42.0%
blood glucose lowering activity in db/db mice model. In vitro anti-hyperglycemic activity evaluation
exhibited that compounds 8a (IC50 = 24.5 lM) and 8b (IC50 = 36.2 lM) are potential PTP-1B inhibitors
Keywords:
thereby revealing their possible mechanism of anti-diabetic action. Compounds 7a, 7b, 8a, 8b, 8d, 8e
and 8f have shown significant anti-dyslipidemic activity in triton induced dyslipidemia in rats.
Ó 2009 Elsevier Ltd. All rights reserved.
Pyranocoumarins
Anti-hyperglycemic
Anti-dyslipidemic
PTP 1B
Diabetes mellitus is multifactorial disease characterized by high
level of blood glucose and impaired insulin action.1 It is an inde-
pendent risk factor for the development of coronary artery dis-
eases, myocardial infarction, hypertension, and dyslipidemia.
Non-insulin dependent diabetes mellitus (NIDDM) accounts for
approximately 80–90% of all diabetes cases.2 The number of dia-
betic complains are continuously growing with a currently esti-
mated worldwide incidence of about 194 million people and
expected to increase to 330 million by 2025.3 When the carbohy-
drates are not present in sufficient amount or their metabolism
is impaired, fats become the principle source of energy. Fatty acids
are mobilized into the general circulation leading to secondary tri-
glyceridemia in which total serum lipids in particular triglycerides
as well as the levels of cholesterol and phospholipids increases.
This rise is proportional to the severity of the diabetes. Uncon-
trolled diabetes is manifested by a very high rise in triglycerides
and fatty acid levels.4 An increase in plasma lipids, particularly
cholesterol, is a common feature of atherosclerosis, a condition
involving arterial damage, which may lead to ischemic heart dis-
ease, myocardial infarction, and cerebro-vascular accidents. These
conditions are responsible for one-third of deaths in industrialized
nations.5 Therefore, an anti-hyperglycemic drug, having lipids (tri-
glycerides and cholesterol) lowering activity is of great demand.
Several research groups have focused their attention to develop
such dual-acting agents.6 Herein, we describe design, synthesis,
anti-hyperglycemic and anti-dyslipidemic activity of pyranocoum-
arin derivatives. The design of pyaranocoumarin is based on natu-
ral products exhibiting promising antidiabetic activity such as
Sanggenone C,7 Luvangetin,8 and Rutamarin.9 These natural prod-
ucts have coumarin as central nucleus along with pyran or furan
scaffolds present in linear arrangements. Based on this we have
designed and synthesized pyranocoumarin having angular
arrangements for their anti-hyperglycemic and lipid lowering
activity (Fig. 1).
The key intermediate 8-acetyl-7-hydroxy-4-methyl-2H-chro-
men-2-one 5 was synthesized from resorcinol 1 in four steps. Resor-
cinol 1 was treated with ethyl acetoacetate 2 and concentrated
H2SO4 to yield 7-hydroxy-4-methyl-2H-chromen-2-one 3. 7-Hydro-
xy-4-methyl-2H-chromen-2-one 3 on reaction with acetic anhy-
dride in pyridine gave 4-methyl-2-oxo-2H-chromen-7-yl acetate
4. Fries migration of the acetyl group of compound 4 by AlCl3 at
170 °C resulted to the formation of 8-acetyl-7-hydroxy-4-methyl-
2H-chromen-2-one 5. Compound 5 was treated with paraformalde-
hyde and amine in different optimized reaction conditions resulting
to the formation of a range of pyranocoumarins. 8-Acetyl-7-hydro-
xy-4-methyl-2H-chromen-2-one 5, when refluxed with 2 equiv of
paraformaldehyde in ethanol afforded 9-ethoxymethyl-4-methyl-
8, 9-dihydro-pyrano[2,3-f]chromene-2,10-dione 6 in 62% yield.
Refluxing 8-acetyl-7-hydroxy-4-methyl-2H-chromen-2-one 5 with
3 equiv of paraformaldehyde and 1 equiv of amine in ethanol affor-
ded compounds 7a–c and 8a–h. 8-Acetyl-7-hydroxy-4-methyl-2H-
chromen-2-one 5, when refluxed with 3 equiv of paraformaldehyde
and 2 equiv of amine in ethanol afforded compounds 9a–h in
* Corresponding author.
E-mail address: dratulsax@gmail.com (A. Kumar).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.09.031

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A. Kumar et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6447–6451
Antidiabetic natural products
Coumarin
Pyran
Coumarin
HO
Coumarin
Furan
O
OH
O
O
OH
O
O
O
O
OH
O
OH
Furan
OAc
O
O
OCH3
HO
Luvangetin
Rutamarin
OH
Sanggenone C
OH
R1O
O
R
O
O
O
Our desigened prototype
Figure 1. Designing of novel pyranocoumarins based on natural antidiabetic scaffolds.
60–75% yields. The systematic representation of pyranocoumarin
synthesis is shown in Scheme 1.
anti-hyperglycemic activity but with mono amine side chains
(7a–c and 8a–h) were showing promising anti-hyperglycemic
activity.
Initially the synthesized compounds 7a–c, 8a–h and 9a–h were
screened in vivo for their anti-hyperglycemic activity in sucrose
loaded model (SLM). Metformin was taken as standard drug in
SLM model which showed 25.0% blood glucose lowering activity
at the dose of 100 mg/kg. In SLM model compounds 7a (28.2%),
7c (30.3%), 8a (35.5%), 8b (38.1%), 8c (31.8%), 8e (30.6%) and 8f
(32.5%) showed blood glucose lowering activity respectively at a
dose of 100 mg/kg body weight (Fig. 2). It was interesting to note
that compound with diamine side chain (9a–h) showed poor
All the compounds which exhibited greater anti-hyperglycemic
activity than metformin in SLM model were further screened in su-
crose challenged streptozotocin induced diabetic rat model (STZ).
In STZ model, compounds 7a (25.3%), 7c (24.0%), 8a (35.5%), 8b
(37.7%), 8c (28.8%), 8e (24.4%) and 8f (18.7%) showed blood glucose
lowering activity respectively at a dose of 100 mg/kg body weight
after 24 h treatment whereas the standard drug metformin
showed 26.6% anti-hyperglycemic activity (Fig. 3).
O
O
Acetic anhydride
conc. H2SO4
rt
+
OC2H5
HO
O
O
HO
OH
Pyridine
H3COCO
OC2H5
O
O
2
3
1
4
H
O
AlCl3
2 eq. paraformaldehyde
O
O
O
Ethanol
reflux
HO
O
O
°
170 C
O
5
6
C2H5O
O
R
O
R
R
HO
R
3 eq. paraformaldehyde
1 eq. amine
O
O
O
O
O
O
O
O
O
O
2 eq. amine
Ethanol
reflux
Ethanol
reflux
8a-h
9a-h
7a-c
N
N
NH,
NH,
NH,
N
R =
NH,
NH,
NH,
NH,
N
NH
O
c
a
d
g
b
f
e
h
Scheme 1. Synthesis of novel pyranocoumarins.

A. Kumar et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6447–6451
6449
Blood glucose lowering activity in SLM model
50
40
30
20
10
0
Compounds
Figure 2. Anti-hyperglycemic activities of compounds 7a–c, 8a–h, and 9a–h in SLM.
Two of the most active compounds in STZ model (8a and 8b)
were further screened for their anti-hyperglycemic activity in db/
db mice which is a well-characterized model of type 2 diabetes.
The background for the db/db mouse is the C57BL/Ks strain. The
major deficiency of the C57BL/KsBom-db mouse (db/db mouse) is
lack of a functional leptin receptor. This leads to defective leptin
signaling and a complete lack of feedback from leptin. Both hypo-
thalamic NPY content and secretion are consequently elevated, and
this result in hyperphagia and decreased energy expenditure, obes-
ity, insulin-resistance, hyperinsulinemia, hyperglycemia and dysli-
pidemia. The optimal age of db/db mice used for experiments is
from week 12 to 18 when they have developed NIDDM with dia-
betic dyslipidemia but still have functional b-cells in the pancreas.
The two compounds exhibited 21.1% (8a), 19.5% (8b) after 5 days
and 38.0% (8a), 42.0% (8b) blood glucose lowering activity after
10 days treatment respectively. The standard drug rosiglitazone
showed 22.0% after 5 days and 48.1% blood glucose lowering activ-
ity after 10 day treatment in similar conditions at a dose of
100 mg/kg (Fig. 4).
Anti-hyperglycemic activity in db/db mice model
60
50
40
30
20
10
0
Day 5
Day 10
8a
8b
Rosiglitazone
Compound
Figure 4. Anti-hyperglycemic activities of compounds 8a, 8b and rosiglitazone in
db/db mice model.
In order to study the possible mechanism of anti-hyperglycemic
activity of these compounds (7a–c, 8a–f and 9a–f), we evaluated
them for their in vitro anti-hyperglycemic activity against the glu-
cose-6-phosphatase, glycogen phosphorylase and
a-glucosidase,
cogen phosphorylase activity is measured by the modified method
of Rall et al.11 Effect of compounds on
a-glucosidase was deter-
DPP IV and PTP-1B enzymes at 100 M concentrations (Table 1).
l
mined by method of Lebovitz12 The dipeptidyl peptidase enzyme
inhibition assays were performed according to the methods of
Wright et al.13 Protein Tyrosine Phosphatase 1B activity was
determined by the modified method14 using pNPP as substrate.
The effect of compounds on glucose-6-phosphatase was studied
by pre-incubating the compound in 1.0 mL reaction system for
10 min and then determining the residual glucose-6-phosphatase
activity according to the method of Hubscher and West.10 The gly-
Anti-hyperglycemic activity of pyranocoumarins in STZ model
35
30
25
20
15
10
5
Metformin
8f
8e
8c
8b
8a
7c
7a
Control
0
-20
0
30
60
90
120
180
240
300
1440
Time (min)
Figure 3. Anti-hyperglycemic activities of compounds 7a, 7c, 8a, 8b, 8c, 8e, and 8f in STZ model.

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A. Kumar et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6447–6451
Table 1
In vitro anti-hyperglycemic activity of compounds 7a–c, 8a–h and 9a–h
Compounds
% Inhibitiona
Glucose-6-phosphatase
Glycogen phosphorylase
a-Glucosidase
DPP IV
PTP 1Bb
7a
7b
7c
8a
8b
8c
8d
8e
8f
8g
8h
9a
9b
9c
9d
9e
3.50
7.36
5.83
12.45
6.56
8.31
24.23
11.10
17.43
31.67
9.67
6.78
13.8
8.28
9.45
20.39
12.96
21.54
28.00
NI
NI
8.3
8.4
8.5
8.3
NI
8.7
34.3
37.3
22.1
NI
NI
12.0
NI
12.0
NI
8.3
NI
12.6
13.2
NI
NI
7.9
NI
83.3 (52.4)
64.0 (62.1)
74.6 (56.7)
92.2 (24.5)
88.5 (36.2)
72.1 (82.7)
74.1 (90.4)
80.5 (40.5)
70.3 (65.5)
62.3 (70.6)
54.5 (76.2)
24.68
33.27
29.78
41.34
43.62
21.98
28.67
41.34
57.8
27.9
NI
26.9
NI
16.2
11.3
8.3
7.3
NI
8.5
NI
6.4
15.9
15.8
—
30.1
12.8
17.2
15.2
12.2
11.1
10.2
22.1
11.0
—
NI
8.8
6.8
8.2
8.1
11.4
12.5
15.2
9.8
—
9f
9g
9h
Sod. ortho-vanadate
a
Compounds were evaluated at 100
IC50 (lM) values are given in parentheses.
l
M concentration; NI means no inhibition.
b
Although the compounds (7a–c, 8a–h, 9a–h) were showing poor
inhibitory activities for glucose-6-phosphatase, glycogen phos-
plasma levels of TC, PL and Tg by 37.1%, 39.1% and 36.1%, respec-
tively at the dose of 100 mg/kg (Fig. 5).
phorylase,
a
-glucosidase and DPP IV enzymes, compounds (7a–c,
Structural activity relationship reveals that compounds having
ether group were showing better antidiabetic activity than corre-
sponding hydroxyl group. Nitrogen containing six member rings
(8a and 8b) were exhibiting more potent antidiabetic activity
in vivo as well as PTP-1B inhibition in comparison to five member
and open chain nitrogen containing compounds. Compound 8b
having piperidine moiety was most potent compound of the series.
It is interesting to note that introduction of nitrogen in piperidine
ring (8f, 8g and 8h) resulted in loss of antidiabetic and PTP 1B
inhibitory activity however incorporation of oxygen in piperidine
ring (8a) have shown similar activity profile as of 8b.
In conclusion, we have synthesized a series of pyranocoumarins
derivatives (7a–c, 8a–h, 9a–h) and evaluated their anti-hypergly-
cemic as well as anti-dyslipidemic activities. Several compounds
of the series have shown promising anti-hyperglycemic activity
in SLM, STZ and db/db mice models. Some compounds of the series
(8a and 8b) were found potential anti-hyperglycemic agents com-
parable to standard drugs. In vitro anti-hyperglycemic activity
8a–h) have shown promising PTP 1B inhibitory activity. It was
interesting to note that compounds with diamine side chain (9a–
h) exhibited moderate whereas compounds with monoamine side
chain (7a–c, 8a–h) exhibited high PTP 1B inhibitory activity. From
these observations, we concluded that PTP 1B might be a possible
target for these anti-hyperglycemic compounds.
The anti-dyslipidemic activities of compounds 7a–c, 8a–h and
9a–h were evaluated in a triton model.15 Administration of triton
WR-1339 in rats (Charles Foster strain, male, adult, body weight
200–225 g) induced marked hyperlipidemia as evidenced by in-
crease in the plasma levels of total cholesterol (TC) (3.50 F), phos-
pholipids (PL) (3.12 F), triglyceride (Tg) (3.14 F). Treatment of
hyperlipidemic rats with compounds (7a–c, 8a–h and 9a–h) at
dose of 100 mg/kg po reversed the plasma level of lipids with vary-
ing extents. Compounds 7a, 7b, 8a, 8b, 8d, 8e and 8f showed signif-
icant lipid lowering activity in plasma level of TC, PL and Tg
respectively, while other compounds showed mild lipid lowering
activity as compared to triton. These data were compared with
Gemfibrozil at a dose of 100 mg/kg, which showed a decrease in
evaluation revealed that compounds 8a (IC50 = 24.5
lM) and 8b
(IC50 = 36.2 M) are potential PTP 1B inhibitors thereby revealing
l
45
40
35
30
25
20
15
10
5
Total cholesterol
Phospholipid
Triglyceride
0
Compounds
Figure 5. Anti-dyslipidemic activities of compounds 7a–c, 8a–h and 9a–h.

A. Kumar et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6447–6451
6451
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