Synthesis of analogues of antihyperglycemic lead karanjin

Article abstract of DOI:10.1007/s00044-010-9387-1

Karanjin isolated in gram scale from Pongamia pinnata was subjected to transformation reactions and 11 analogues were prepared and evaluated for their antihyperglycemic activity in streptozotocin-induced diabetic rats, and compared with the standard drug metformin at the dose of 100 mg/kg body weight. Four derivatives showed comparable activity with metformin. Springer Science+Business Media, LLC 2010.

Full text of DOI:10.1007/s00044-010-9387-1

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2011) 20:1465–1472
DOI 10.1007/s00044-010-9387-1
ORIGINAL RESEARCH
Synthesis of analogues of antihyperglycemic lead karanjin
•
•
•
Akanksha Prem P. Yadav Arvind K. Srivastava
Rakesh Maurya
Received: 15 March 2010 / Accepted: 5 July 2010 / Published online: 18 July 2010
Ó Springer Science+Business Media, LLC 2010
Abstract Karanjin isolated in gram scale from Pongamia
pinnata was subjected to transformation reactions and 11
analogues were prepared and evaluated for their antihy-
perglycemic activity in streptozotocin-induced diabetic
rats, and compared with the standard drug metformin at the
dose of 100 mg/kg body weight. Four derivatives showed
comparable activity with metformin.
karanjin was the first representative of the furanoflavonol
class of the compounds, which were found restricted to
some leguminous plant species.
Major source for karanjin are Pongamia and Tephrosia
species, it occurs as a major component of the plant. Pre-
viously, we have reported karanjin and pongamol as the
lead compounds with antihyperglycemic activity from
chloroform fraction of the ethanolic extract of the Pong-
amia pinnata fruits (Yadav et al., 2004; Tamrakar et al.,
2008). Karanjin possessed hypoglycemic (Mandal and
Maity, 1986), CNS stimulant (Mahli et al., 1989) and
nitrification inhibitor (Majumdar, 2008) activities. It has
been reported that the non lipid fraction of the Pongamia
oil containing karanjin and pongamol was responsible for
haemotoxicity present in the oil (Gandhi and Cherian,
2000). As per the synthetic studies are concerned it has not
been explored much because of the complications regard-
ing generation of angular furan ring, which was peculiarity
of these compounds. Karanjic acid, one of the degradation
products has been utilized for the synthesis of karanjin and
its analogues most of the time and no synthetic transfor-
mation as such have been done previously (Ahmad et al.,
2006). In order to study the impact of different substituents
on antihyperglycemic activity, we have performed deriva-
tization at flavonol hydroxyl of karanjin, which is isolated
in gram scale from Pongamia pinnata fruits.
Keywords Karanjin ꢀ Antihyperglycemic activity ꢀ
STZ ꢀ Metformin
Introduction
Diabetes mellitus is a group of syndrome characterized by
hyperglycemia (altered metabolism of lipid, carbohydrates
and proteins) and an increased risk of complications from
vascular disease. Owing to several side-effects and toxicity
affiliated with modern antidiabetic drugs, newer targets and
agents merit a serious attention (Oh et al., 2005). Natural
products have been important source for new chemical
entities having great deal regarding their therapeutic
potentials even to the most threatening ailments to the
human kind (Nisbet and Moore, 1997). The natural product
CDRI communication no. 7944.
Results and discussion
Akanksha ꢀ P. P. Yadav ꢀ R. Maurya (&)
Medicinal and Process Chemistry Division, Central Drug
Research Institute, CSIR, Lucknow 226 001, India
e-mail: mauryarakesh@rediffmail.com
Chemistry
Demethylation of karanjin to karanjonol (1) (Gupta et al.,
1982) was achieved by treating karanjin with boron tri-
bromide (Scheme 1).
A. K. Srivastava
Biochemistry Division, Central Drug Research Institute,
CSIR, Lucknow 226 001, India
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Scheme 1 Synthesis of
karanjonol (1)
O
BBr3, dry DCM, -780
O
O
O
OH
OCH3
O
O
1
Karanjin
O
O
O
O
O
O
NaOMe
1'''
O
OH
O
O
9
1'''
OAc
O
O
O
O
HO
8
OH
OH
1'''
3'''
AcO
O
2'''
OAc
OAc
2
Br
O
O
O
O
K2CO3
O
O
O
OH
2'''
1'''
O
Phenacyl
bromide
Br
O
O
O
1
1'''
O
3'''
7
O
3
4'''
Cl
O
O
O
O
O
O
Silica gel
2'''
1'''
O
OH
O
3'''
6'''
1'''
2'''
O
O
O
N
1'''
7'''
3'''
2'''
O
HO
O
OH
4'''
3'''
5
4
5'''
6
Scheme 2 Synthesis of analogues of karanjin
Demethylkaranjin, a natural product known as karanjo-
nol (1) was further subjected to chemical transformation
based on enolate chemistry. As shown in Scheme 2 the
enolate generated from karanjonol (1) was further reacted
with the alkyl halides such as allyl bromide, isopropyl
bromide, epichlorohydrin and phenacyl bromide to produce
the corresponding 3-O-substituted furanoflavonols 2–4 and
7, respectively. Products obtained were purified by column
chromatography except compound 4. Epoxide group pres-
ent in the compound 4 was labile enough to be cleaved by
the silica gel, afforded a hygroscopic gummy compound 5,
which was identified as the hydrolysed product of the
corresponding epoxide through NMR, two methylene car-
bons appeared at d 74.9 and 63.7 and methine carbon
appeared at d 70.9, also supported by FAB MS spectros-
copy [M ? H]? at m/z 353. 3-Oxiranylmethoxyfurano-
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Med Chem Res (2011) 20:1465–1472
1467
Table 1 Antidiabetic activity of analogues of Karanjin
O
O
O
O
Compound
% Activity STZ-S model
5 h
O
H2, Pd/C
24 h
OCH3
OCH3
Karanjin
-30.8*
-37.2**
-33.0*
-26.9*
-4.8
-42.6***
-39.0**
-16.6
-15.4
-37.0**
?22.9
?0.5
O
10
Karanjin
1
BBr3,
dry DCM
2
3
6
7
?15.4
?9.7
O
O
O
O
H2, Pd/C
8
10
?6.2
?9.8
OH
OH
11
?7.0
?5.3
O
O
11
Metformin
-26.7*
25.0*
1
* P \ 0.05, ** P \ 0.01,*** P \ 0.001
Scheme 3 Synthesis of 200, 300-dihydrokaranjonol (11)
flavone (4) was reacted with diethyl amine to produce the
corresponding 3-(3-diethylamino-2-hydroxy-propoxy)-fur-
anoflavone (6).
5 h. Substitution of 3-OH in compound 1 with allyl bromide,
isopropyl bromide and 3-diethylamino-2-hydroxy-propoxy
groups showed promising activity by 33.0%, 26.9% (5 h)
and 37.0% (24 h), respectively, suggesting that these groups
at 3-position contribute to the antihyperglycemic activity of
the respective compounds and can serve as good leads for
further synthetic studies. Other transformed compounds
showed insignificant or no activity.
Glycosides of the furanoflavonoids are very rare, as only
three furanoflavonoid glycosides have been reported
(Ahmad et al., 2004). Different glycosyl donors are
reported for the purpose of glycosidation in the literature.
As per the reported methods for the glycosidation of
flavonols at 3-position (Li et al., 2002), we opted for the
tetraacetylglucopyranosyl bromide as glycosyl donor for
the acceptor, enolate of karanjonol. This is the first report
for synthesis of the glycosides of furanoflavonoids. Kara-
njonol on reaction with tetraacetylglucopyranosyl bromide
produce 3-O-tetraacetylglucoside of karanjin (Scheme 2).
Deacetylation afforded 3-O-glucosylkaranjonol (9). Ana-
logues 3–9 are new, synthesized for the first time and are
characterized from their spectroscopic studies.
Conclusion
We have prepared 11 analogues of antidiabetic lead com-
pound karanjin. As there was only one possible centre to
play around, i.e., flavonol hydroxyl group, we have
developed our transformation strategies accordingly and
synthesized analogues for karanjin to evaluate their anti-
diabetic activity. Transformed compounds were tested for
their antidiabetic activity in vivo in streptozotocin (STZ)
induced b-cell damaged diabetic model of Sprague–
Dawley strain male albino rats. Biological studies revealed
that compounds 1, 2, 3 and 6 have comparable activity with
karanjin. Compound 6 can be further derivatized to
enhance the activity. Other transformed compounds have
shown insignificant or no activity.
When karanjin was hydrogenated in the Parr assembly at
50 psi at rt, produced 200, 300-dihydrokaranjin (10). Com-
pound 10 was demethylated with BBr3 to obtain the cor-
responding demethylated product 200, 300-dihydrokaranjonol
(11). Further, compound 11 was also prepared from com-
pound 1 by hydrogenation under medium pressure (50 lbs/
sq. inch) as shown in Scheme 3.
Biological activity of the analogues
Karanjin, compounds 1–3, 6–8, 10 and 11 were tested for
their antidiabetic activity in vivo in streptozotocin (STZ)
induced b-cell damaged diabetic model of Sprague–Dawley
strain male albino rats at the dose of 100 mg/kg p.o. taking
metformin as a standard. The antidiabetic activities of
compounds are mentioned in the Table 1. Biological studies
revealed that compounds 1, 2, 3 and 6 showed significant
activity. From the results, it is evident that demethylated
product (compound 1) showed increased activity (37.2%) at
Experimental procedure
Detection of purity of isolated compound was checked by the
use of ultraviolet fluorescent lamp have the wavelength k 254
(short wavelength) and by spraying of 10% sulphuric acid in
methanol and heating the TLC plate on hot plate. TLC was
run on pre-coated silicagel 60 F254 (Merck). An IR spectrum
was recorded on Perkin-Elmer RX-1 spectrophotometer
using KBr pallets. The ESI Mass spectra were recorded using
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Med Chem Res (2011) 20:1465–1472
a beam of Argon (2–8 eV) on Joel SX102/DA 6000 Mass
spectrometer. The NMR spectra were recorded in CDCl3 at
300 MHz on an Advance DPX 200 spectrometer; chemical
shifts are reported in d ppm downfield from TMS, used as
internal standard. Silica gel (60–120 mesh) was used for
column chromatography. In the case of flash chromatogra-
phy, mesh size of silica gel was (230–400 mesh).
8.17 (1H, d, J = 8.8 Hz, H-5), 7.78 (1H, d, J = 1.94 Hz,
H-200), 7.48–7.59 (4H, m, H-30, 40, 50, 6), 7.25 (1H, brd,
J = 1.08 Hz, H-300); 13C NMR (CDCl3, 50 MHz) d: 173.7
(C-4), 158.5 (C-7), 150.7 (C-9), 146.1 (C-200), 144.4 (C-2),
138.9 (C-3), 131.5 (C-10), 130.4 (C-40), 129.0 (C-30, 50),
127.8 (C-20, 60), 121.9 (C-5), 117.5 (C-10), 116.4 (C-8),
110.5 (C-6), 104.6 (C-300); FAB MS (pos.): m/z 279
[M ? H]?, 557 [2 M ? H]?.
Extraction of fruits of Pongamia pinnata
General procedure for generation of potassium enolate
of karanjonol (1)
Air-dried and powdered fruits of Pongamia pinnata (6 kg)
were extracted with ethanol (10 L) at room temperature. The
ethanolic extract (750 g) was triturated successively with
n-hexane (1 l 9 20) and chloroform (500 ml 9 15) and
soluble fractions were concentrated under reduced pressure
to obtain n-hexane fraction (360 g) and chloroform fraction
(70 g). On activity-guided fractionation, the chloroform
fraction was subjected to column chromatography over flash
silica gel (230–400 mesh) eluting with a gradient of C6H6–
EtOAc (1:0–1:1) to afford 60 fractions. These fractions were
pooled into nine sub-fractions (F-1 to F-9) according to their
TLC pattern. Flash CC of F-2 using C6H6 as eluent yielded
karanjin. The structure elucidation of the compound was
performed by spectroscopic techniques, as reported in our
previous communication (Yadav et al., 2004).
Under nitrogen atmosphere, anhydrous K2CO3 (1.50 g)
was placed in a 50 ml RB and added karanjonol (1)
(200 mg, 0.72 mmol) dissolved in dry acetone (15 ml).
Reaction mixture was stirred for half an hour for generation
of potassium enolate of karanjonol as indicated by yellow
colouration of reaction mixture and then alkylated with
corresponding alkyl bromide.
Procedure for synthesis of 3-O-allylkaranjonol (2)
To the enolate of karanjonol (1) (200 mg, 0.72 mmol)
under nitrogen, added drop wise allyl bromide (130 mg and
1.08 mmol) and stirred the reaction mixture for 4 h at room
temperature. Reaction mixture was filtered through celite
and concentrated the filtrate to give crude product which
was purified by column chromatography over silica gel
(60–120 mesh) using isocratic elution with hexane:ethyl
acetate (9:1), afforded 2, yield 195 mg (85%); white
crystals, mp 145–146°C; IR (KBr) mmax: 2943, 1628, 1586,
1458, 1410, 1348, 1202, 1163, 1039, 932, 759 and
693 cm-1; UV (CHCl3) kmax: 261, 302 nm; 1H NMR
(CDCl3, 200 MHz) d: 8.16 (3H, m, H-20, 60, H-5), 7.75
(1H, d, J = 2.1 Hz, H-200), 7.51–7.56 (4H, m, H-30, 40, 50
and H-6), 7.17 (1H, brd, J = 2.05, 1.15 Hz, H-300), 5.94
(1H, m, H-2000), 5.12-5.33 (2H, m, H-3000), 4.68 (2H, d,
J = 6.0 H-1000); 13C NMR (CDCl3, 50 MHz) d: 175.4
(C-4), 158.5 (C-7), 155.6 (C-2),150.3 (C-9), 146.1 (C-200),
140.7 (C-3), 134.0 (C-2000), 131.5 (C-10), 130.9 (C-40),
129.0 (C-30, 50), 128.8 (C-20, 60), 120.0 (C-8), 122.2 (C-5),
118.9 (C-3000), 110.3 (C-6), 104.6 (C-300), 73.7 (C-10), 117.4
(C-10); FAB MS (pos.): m/z 319 [M ? H]?, 637
[2 M ? H]?. Elemental analysis: calc. for C20H14O4:C,
75.46; H, 4.43; found: C, 75.51, H, 4.39%.
Procedures for transformation of karanjin
Procedure for demethylation of karanjin to afford
karanjonol (1)
In a 100 ml RB flask, 2.00 g (6.8 mmol) of karanjin dis-
solved in 50 ml of dry dichloromethane was placed under
nitrogen atmosphere. Maintained the temperature at –78°
and added 27.3 ml (4 eq) of BBr3 (1 M solution in DCM)
drop wise with stirring. Stirring was continued for 3 h at
–78° and then warmed to room temperature and stirred for
1 h at room temperature. After completion of the reaction,
the reaction mixture was quenched by drop wise addition
of water and poured the reaction mixture into the saturated
solution of sodium bicarbonate (50 ml). Reaction mixture
was extracted with dichloromethane (50 ml 9 5), washed
with brine (50 ml 9 3), dried over anhydrous sodium
sulphate and concentrated to get crude product. Crude
product was purified by column chromatography over silica
gel (60–120 mesh) by isocratic elution with benzene as
eluent to afford karanjonol (1), yield 1.7 g (90%); white
crystals, mp 192–193°C; IR (KBr) mmax: 3467, 2924, 1622,
1586, 1471, 1389, 1292, 1161, 1117, 1041, 754 and
681 cm-1; UV (CHCl3) kmax: 261, 302 nm; 1H NMR
(CDCl3, 200 MHz) d: 8.29 (2H, brdd, J = 6.9 Hz, H-20, 60),
Procedure for synthesis of 3-O-isopropylkaranjonol (3)
Potassium enolate of karanjonol (1) (250 mg, 0.89 mmol)
was generated under nitrogen added drop wise isopropyl
bromide (164 mg, 1.33 mmol) and refluxed the reaction
mixture for 4 h. Reaction mixture was filtered through
celite and concentrated the filtrate to give crude product
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1469
which was purified by column chromatography over silica
gel (60–120 mesh) using isocratic elution with hexane:
ethyl acetate (9:1) to afford 3; yield 205 mg (71%); white
needles, mp 156–157°C; IR (KBr) mmax: 2972, 1631, 1464,
1411, 1375, 1285, 1217, 1166, 1046 and 758 cm-1; UV
3-(2,3-dihydroxy-propoxy)-karanjonol (5): gummy; IR
(KBr) mmax: 3508, 2930, 1638, 1606, 1465, 1279, 1245,
1168, 1033, 944 and 769 cm-1; UV (CHCl3) kmax: 266,
301 nm; 1H NMR (CDCl3, 200 MHz) d: 8.11–8.21 (3H, m,
H-20, 60 and H-5), 7.78 (1H, d, J = 2.1 Hz, H-200),
7.56–7.60 (4H, m, H-30, 40, 50 and H-6), 7.19 (1H, brd,
J = 1.3 Hz, H-300), 3.97 (3H, m, H-2000, 3000), 3.91 (2H, brs,
H-1000); 13C NMR (CDCl3, 50 MHz) d: 176.1 (C-4), 158.7
(C-7), 156.4 (C-9), 150.5 (C-2), 146.3 (C-200), 141.0 (C-3),
131.5 (C-40), 130.8 (C-10), 129.2 (C-30, 50), 128.7 (C-20, 60),
122.2 (C-5), 119.4 (C-8), 117.4 (C-9), 110.8 (C-6), 104.5
(C-300), 74.9 (C-1000), 70.9 (C-2000), 63.7 (C-3000); FAB MS
(pos.): m/z 353 [M ? H]?, 705 [2 M ? H]?.
1
(CHCl3) kmax: 260, 303 nm; H NMR (CDCl3, 200 MHz)
d: 8.20 (3H, m, H-20, 60, H-5), 7.75 (1H, d, J = 1.9 Hz,
H-2), 7.50-7.57 (4H, m, H-30, 40, 50 and H-6), 7.18 (1H, d,
J = 1.34 Hz, H-300), 4.73 (1H, sept, J = 6.2 Hz, H-1000),
1.21 (6H, d, J = 6.2 Hz, H-2000, 3000); 13C NMR (CDCl3,
50 MHz) d: 175.9 (C-4), 158.5 (C-7), 155.8 (C-2),150.3
(C-9), 146.0 (C-200), 140.1 (C-3), 132.0 (C-10), 130.7
(C-40), 129.2 (C-30, 50), 128.6 (C-20, 60), 122.3 (C-5), 120.0
(C-8), 117.3 (C-10), 110.2 (C-6), 104.6 (C-300), 75.3
(C-1000), 22.8 (C-2000, 3000); FAB MS (pos.): m/z 321
[M ? H]?, 641[2 M ? H]?. Elemental analysis: calc. for
C20H16O4: C, 74.99; H, 5.03; found: C, 75.44; H, 4.85%.
Procedure for synthesis of 3-(3-diethylamino-2-
hydroxy-propyl)-karanjonol (6)
3-O-(2,3-oxiranylmethyl)-karanjonol (4) (250 mg, 0.74 mmol)
and diethyl amine (162 mg and 2.22 mmol) were dissolved
in ethanol (15 ml) and refluxed on an oil bath for 10 h.
Ethanol was evaporated from the reaction mixture and
product was dissolved in water and extracted with ethyl
acetate. Ethyl acetate extract was dried over sodium sul-
phate and concentrated to give crude product, which was
subjected to column chromatography over silica gel
(60–120 mesh) using gradient elution with hexane:ethyl
acetate (9:1–7:3) to afford 6 as brown viscous compound;
yield 140 mg (46%); viscous; IR (KBr) mmax: 3510, 2945,
1633, 1593, 1465, 1353, 1235, 1168, 1015, 945 and
760 cm-1; UV (CHCl3) kmax: 265, 303 nm; 1H NMR
(CDCl3, 200 MHz) d: 8.14 (3H, m, H-20, 60 and H-5), 7.76
(1H, d, J = 1.7 Hz, H-200), 7.52-7.56 (4H, m, H-30, 40, 50 and
H-6), 7.16 (1H, brs, H-300), 4.18 (1H, m, H-2000), 4.06 (2H,
brs, H-1000), 2.82–3.07 (6H, m, H-3000, 4000, 6000), 1.18 (6H, t,
J = 7.0 Hz, CH3-5000, 7000); 13C NMR (CDCl3, 50 MHz) d:
175.2 (C-4), 158.1 (C-7), 155.4 (C-2), 149.9 (C-9), 145.8
(C-200), 140.6 (C-3), 130.8 (C-40), 130.5 (C-10), 128.6 (C-30,
50), 128.4 (C-20, 60), 121.6 (C-5), 119.2 (C-8), 116.9 (C-10),
110.1 (C-6), 104.1 (C-300), 75.1 (C-1000), 66.4 (C-2000), 56.1
(C-3000), 47.7 (C-4000, 6000), 10.4 (CH3-5000, 7000); FAB MS
(pos.): m/z 408 [M ? H]?, 815 [2 M ? H]?. Elemental
analysis: calc. for C24H25O5N: C, 70.74; H, 6.18; N, 3.44;
found: C, 70.61; H, 5.97; N, 3.69%.
Procedure for synthesis of 3-O-(2, 3-oxiranylmethyl)-
karanjonol (4) and 3-(2, 3-dihydroxy-propoxy)-
karanjonol (5)
To the potassium enolate of karanjonol (1, 250 mg,
0.89 mmol) under nitrogen added drop wise epichlorohy-
drin (166 mg, 1.78 mmol) and refluxed the reaction mix-
ture for 8 h. After completion of reaction, the reaction
mixture was filtered through celite and concentrated the
filtrate to give crude product which was purified by column
chromatography over silica gel (60–120 mesh) using iso-
cratic elution with hexane: ethyl acetate (22:3) to afford 5
(hydrolysed product of epoxide ring); yield 155 mg (49%).
Same reaction was performed again with the same
equivalents of reactants and reagents and extracted same as
before but column chromatography of product was per-
formed over deactivated silica gel (deactivated by addition
of 12% of water) eluting with the solvent system hexane:
ethyl acetate (9:1). Recovery of the product was most
efficient in the case of deactivated silica gel as adsorbent
with yield of 60% of 4.
3-O-(2, 3-oxiranylmethyl)-karanjonol (4): viscous; IR
(KBr) mmax: 2928, 1630, 1593, 1465,1289, 1217, 1168, 1042
1
and 769 cm-1; UV (CHCl3) kmax: 261, 307 nm; H NMR
(CDCl3, 200 MHz) d: 8.11–8.20 (3H, m, H-20, 60 and H-5),
7.78 (1H, d, J = 2.1 Hz, H-200), 7.60 (4H, m, H-30, 40, 50 and
H-6), 7.19 (1H, brd, J = 1.3 Hz, H-300), 4.18 (1H, m, H-2000),
4.07 (2H, m, H-1000), 3.70 (2H, m, H-3000); 13C NMR (CDCl3,
50 MHz) d: 176.1 (C-4), 158.8 (C-7), 156.8 (C-9), 146.3
(C-2), 146.3 (C-200), 141.0 (C-3), 131.6 (C-40), 130.8 (C-10),
129.3 (C-30, 50), 128.8 (C-20, 60), 122.2 (C-5), 119.4 (C-8),
117.4 (C-10), 110.9 (C-6), 104.5 (C-300), 75.0 (C-1000), 70.8
(C-2000), 44.5 (C-3000); FAB MS (pos.): m/z 335 [M ? H]?,
669 [2 M ? H]?. Elemental analysis: calc. for C20H14O5:
C, 71.85; H, 4.22; found: C, 70.99, H, 4.32%.
Procedure for synthesis of 3-O-(2-oxo-2-phenyl-ethyl)-
karanjonol (7)
Potassium enolate karanjonol (1) (250 mg, 0.89 mmol)
was generated, under nitrogen, added drop wise phenacyl
bromide (265 mg, 1.33 mmol) in dry acetone (10 ml), the
reaction mixture was stirred for 4 h at room temperature.
After completion of reaction, the reaction mixture was
filtered through celite and concentrated the filtrate to afford
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Med Chem Res (2011) 20:1465–1472
crude product which was purified by column chromatog-
raphy over silica gel (60–120 mesh) using isocratic elution
with hexane:ethyl acetate (9:1) to afford 7; yield 255 mg
concentrated the filtrate to give crude product which was
purified by column chromatography over silica gel (60–120
mesh) using isocratic elution with hexane: ethyl acetate
(88:12) to afford compound 8; yield 270 mg (62%); white
crystals, mp 148–149°C; IR (KBr) mmax: 2938, 1755, 1629,
(72%); white crystals, mp 204–205°C; IR (KBr) mmax
:
2907, 1701, 1630, 1442, 1364, 1239, 1215, 1167, 959 and
757 cm-1; UV (CHCl3) kmax: 255, 305 nm; 1H NMR
(CDCl3, 200 MHz) d: 8.23 (2H, m, H-20, 60), 8.17 (1H, d,
J = 8.8 Hz, H-5), 7.98 (2H, brd, J = 7.8 Hz, H-300, 800),
7.77 (1H, d, J = 2.1 Hz, H-200), 7.44–7.58 (7H, m, H-30, 40,
50, 3000, 4000, 5000 and H-6), 7.23 (1H, d, J = 1.95 Hz, H-300),
5.62 (2H, s, -OCH2CO–); 13C NMR (CDCl3, 50 MHz) d:
194.6 (–OCH2CO–), 175.1 (C-4), 158.6 (C-7), 154.9 (C-2),
150.2 (C-9), 146.1 (C-200), 140.4 (C-3), 135.1 (C-1000),
133.9 (C-4000), 131.1 (C-10), 131.1 (C-40), 129.1 (C-2000,
6000), 129.0 (C-30, 50), 128.9 (C-3000, 5000), 128.5 (C-20, 60),
122.1 (C-5), 119.8 (C-8), 117.4 (C-10), 110.5 (C-6), 104.6
(C-300), 74.1 (–OCH2CO–); FAB MS (pos.): m/z 397
[M ? H]?, 792 [2 M ? H]?. Elemental analysis: calc. for
C25H16O5: C, 75.75; H, 4.07; found: C, 75.87, H, 3.95%.
1599, 1463, 1373, 1223, 1168, 1040, 905 and 762 cm-1
;
UV (CHCl3) kmax: 260, 301 nm; 1H NMR (CDCl3,
200 MHz) d: 8.10 (3H, m, H-20, 60 and H-5), 7.75 (1H, d,
J = 1.8 Hz, H-200), 7.52–7.56 (4H, m, H-30, 40, 50 and 6),
7.13 (1H, brs, H-300), 5.77 (1H, d, J = 7.3 Hz, H-1000), 5.27
(2H, m, H-2000, 4000), 5.08 (1H, t, J = 9.5, 9.1 Hz, H-3000),
3.98 (2H, brs, H-6000), 3.71 (1H, brd, J = 9.8 Hz, H-5000),
1.88, 1.99, 2.01 and 2.13 (3H each, s, COCH3); 13C NMR
(CDCl3, 50 MHz) d: 174.2 (C-4), 170.6-169.8 (49 CO
CH3), 158.5 (C-7), 156.8 (C-2), 150.3 (C-9), 146.2 (C-200),
137.0 (C-3), 131.1 (C-10, C-40), 129.4 (C-30, 50), 128.6 (C-
20, 60), 121.9 (C-5), 119.9 (C-8), 117.4 (C-10), 110.5 (C-6),
104.6 (C-300), 99.3 (C-1000), 73.2 (C-5000), 72.0 (C-2000, C-3000),
68.8 (C-4000), 61.9 (C-6000), 20.8, 20.9, 21.2, 23.0 (49
COCH3); FAB MS (pos.): m/z 609 [M ? H]?, 279 [M?-
glu]?, 331 [glu]?. Elemental analysis: calc. for C31H28O13:
C, 61.18; H, 4.64; found: C, 60.98, H, 4.77%.
Procedure for synthesis of tetre-O-acety-
a-D-glucopyranosyl bromide
Procedure for deacylation of 3-O-
tetraacetylglucopyranosylkaranjonol (8) to produce
3-O-glucopyranosylkaranjonol (9)
a-D-Glucose monohydrate (5.50 g) was added to a mixture
of acetic anhydride (20 ml) and perchloric acid (0.12 ml,
70%) at 30–40° during the course of 1 h. After addition of
red phosphorous (0.15 g), the reaction mixture was cooled
in ice-salt bath; added bromine (2.9 ml) drop wise at such a
rate as to keep the internal temperature below 20°. Sub-
sequently, water (1.5 ml) was added during the course of
10 min., with careful control of the temperature. The vessel
was closed and kept for 3 h at room temperature. The
mixture was diluted with dichloromethane and filtered
through a glass funnel plugged with glass wool. The filtrate
was transferred to separating funnel and washed with ice-
cooled water, saturated sodium bicarbonate solution (vig-
orous evolution of CO2) and finally filtered through a bed
of silica gel and concentrated the filtrate under reduced
pressure to give tetre-O-acety-a-D-glucopyranosyl bromide
which was further purified by crystallization from hexane;
mp 88–89°, yield 90%.
To the stirred solution of sodium methoxide (14 mg,
0.41 mmol) in methanol (5 ml) under nitrogen added 3-O-
tetraacetylglucopyranosylkaranjonol (8) (250 mg, 0.41 mmol)
in dry methanol (25 ml). The reaction mixture stirred at
room temperature for 1 h, then added sufficient amount of
ion exchange resin IR 120 (H?) to render the solution
neutral to pH paper. Removed the resin by filtration,
washed with methanol and evaporated the combined filtrate
under reduced pressure to afford 9 (149 mg, 82%); white
crystals, mp 228–229°C; IR (KBr) mmax: 3460, 2945, 1639,
1599, 1458, 1373, 1213, 1174, 1035, 920 and 755 cm-1
;
UV (CHCl3) kmax: 256, 307 nm; 1H NMR (CDCl3,
200 MHz) d:8.33 (2H, m, H-20, 60), 8.27 (1H, d, J =
1.9 Hz, H-200), 8.06 (1H, d, J = 8.8 Hz, H-5), 7.80 (1H, d,
J = 8.8 Hz, H-6), 7.59 (3H, m, H-30, 40, 50), 7.49 (1H, d,
J = 1.48 Hz, H-300), 5.65 (1H, d, J = 7.09 Hz, H-1000),
3.12–3.73 (6H, m, H-2000, 3000, 4000,5000, 6000); 13C NMR
(CDCl3, 50 MHz) d: 173.9 (C-4), 157.8 (C-7), 155.3 (C-2),
149.6 (C-9), 147.8 (C-200), 137.1 (C-3), 131.0 (C-10, C-40),
129.4 (C-30, 50), 128.6 (C-20, 60), 121.5 (C-5), 119.2 (C-8),
117.1 (C-10), 110.6 (C-6), 104.7 (C-300), 101.1 (C-1000),
77.9 (C-5000), 76.8 (C-3000), 74.5 (C-2000), 70.2 (C-4000), 61.1
(C-6000); FAB MS (pos.): m/z 441 [M ? H]?, 881
[2 M ? H]?. Elemental analysis: calc. for C23H20O9: C,
62.73; H, 4.58; found: C, 62.59, H, 4.63%.
Procedure for synthesis of 3-O-
tetraacetylglucopyranosyl karanjonol (8)
Potassium enolate of karanjonol (1) (200 mg, 0.72 mmol)
was generated under nitrogen added drop wise solution of
tetre-O-acety-a-D-glucopyranosyl bromide in dry acetone
(591 mg, 1.44 mmol), stirred the reaction mixture for 4 h
at room temperature. After completion of reaction,
the reaction mixture was filtered through celite and
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Med Chem Res (2011) 20:1465–1472
1471
Procedure for synthesis of 200, 300-dihydrokaranjin (10)
kept for overnight starvation and were made diabetic by
injecting streptozotocin (STZ) intraperitonially at a dose
of 60 mg/kg, prepared in 100 mM citrate buffer (pH 4.5).
Fasting blood glucose level was measured by using Glu-
cometer (ACCU-CHEK II; Roche Diagnostics, USA).
After 48 h, animals showing blood glucose level above
10 mM (180 mg/dl) were considered as diabetic. The
diabetic rats with fasting blood glucose values from 10 to
18 mM (180–324 mg/dl) were included in this study.
Animals were divided into groups consisting of five ani-
mals in each. Animals of one group were regarded as
control group (orally administered 1% gum acacia) and
other groups were treated as experimental groups. Rats of
experimental groups were given suspension of the test
sample made in 1% gum acacia. Animals of control group
were given an equal amount of vehicle (1% gum acacia).
An oral sucrose load of 2.5 g/kg was given to all groups,
30 min post administration of the test sample/vehicle.
Blood glucose levels of the animals of all groups were
again measured at 30, 60, 90, 120, 180 and 300 min and
24 h post sucrose load. Food (not water) was removed
from the cages during the experimental period. Percent
hyperglycemic activity was determined by comparing the
AUC (Area Under Curve) of experimental groups with
that of vehicle-treated control group.
Karanjin (200 mg, 0.68 mmol) was hydrogenated by 10%
palladium over charcoal in ethanol at NTP no reaction
observed, and then the same reaction mixture was trans-
ferred to the Parr assembly at 50 psi at rt, reaction com-
pleted in 8 h. Reaction mixture was filtered through Celite
and further purification by column chromatogrphy over
silica gel (60–120 mesh) using isocratic elution with ben-
zene to afford 200, 300-dihydrokaranjin (10, 191 mg) in 95%
yield; white crystals, mp 186–187°C; IR (KBr) mmax: 2974,
1628, 1601, 1445, 1388, 1226, 1163, 1028, 953 and
771 cm-1; UV (CHCl3) kmax: 258, 315 nm; 1H NMR
(CDCl3, 200 MHz) d: 8.04–8.12 (3H, m, H-20, 60, H-5),
7.49–7.52 (3H, m, H-30, 40, 50), 6.88 (1H, d, J = 8.6 Hz,
H-6), 4.80 (2H, t, J = 8.9 Hz, H-200), 3.45 (2H, t, J =
8.8 Hz, H-300), 3.88 (3H, s, OCH3); 13C NMR (CDCl3,
50 MHz) d: 175.0 (C-4), 165.8 (C-7), 154.8 (C-2),153.1
(C-9), 141.6 (C-3), 131.5 (C-10), 130.9 (C-40), 128.9 (C-30,
50), 128.6 (C-20, 60), 127.8 (C-5), 118.9 (C-8), 113.5 (C-10),
108.9 (C-6), 73.3 (C-200), 60.5 (–OCH3), 27.0 (C-300); FAB
MS (pos.): m/z 295 [M ? H]?, 589 [2 M ? H]?. Ele-
mental analysis: calc. for C18H14O4: C, 73.46; H, 4.79;
found: C, 73.76, H, 4.58%.
Procedure for synthesis of 200, 300-dihydrokaranjonol (11)
Acknowledgements Akanksha would like to thank CSIR, New
Delhi for providing Senior Research Fellowship. SAIF, CDRI is
acknowledged for providing spectroscopic data.
Procedure for demethylation of karanjin; was repeated with
compound 10 (250 mg, 0.85 mmol), 3.4 ml (4 eq) of BBr3
(1 M solution in DCM) and crude product was purified by
column chromatography over silica gel (60–120 mesh) by
isocratic elution with benzene as eluent to afford 200, 300-
dihydrokaranjonol (11), yield 210 mg (88%); white crys-
tals, mp 190–191°; IR mmax(KBr) cm-1: 3430, 2969, 1622,
1596, 1470, 1380, 1339, 1282, 1163, 1060, 769 and 670;
UV kmax CHCl3: 258, 315; 1H NMR (CDCl3, 200 MHz) d:
8.25 (2H, m, H-20, 60), 8.07 (1H, d, J = 8.6 Hz, H-5),
7.41–7.56 (3H, m, H-30, 40, 50), 6.91 (1H, d, J = 8.6 Hz,
H-6), 4.83 (2H, t, J = 8.9 Hz, H-200), 3.50 (2H, t,
J = 8.9 Hz, H-300); 13C NMR (CDCl3, 50 MHz) d: 174.6
(C-4), 163.4 (C-7), 154.8 (C-2),152.0 (C-9), 139.2 (C-3),
131.5 (C-10), 130.9 (C-40), 128.6 (C-30, 50), 128.3 (C-20, 60),
126.9 (C-5), 117.7 (C-8), 113.5 (C-10), 109.2 (C-6), 73.5
(C-200), 27.3 (C-300); FAB MS (pos.): m/z 281 [M ? H]?,
561 [2 M ? H]?. Elemental analysis: calc. for C17H12O4:
C, 72.85; H, 4.32; found: C, 73.03, H, 4.14%.
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