Synthesis of chromenochalcones and evaluation of their in vitro antileishmanial activity

Article abstract of DOI:10.1016/j.bmc.2005.07.005

A large number of novel chromenochalcones were synthesized by pyridine-catalysed chromenylation of mono-chelated meta-dihydric acetophenones with the monoterpene, citral dimethyl acetal and subsequent Claisen-Schmidt condensation of the resultant acylchro

Full text of DOI:10.1016/j.bmc.2005.07.005

Bioorganic & Medicinal Chemistry 13 (2005) 6543–6550
Synthesis of chromenochalcones and evaluation of their in vitro
antileishmanial activityq
Tadigoppula Narender,a,* Tanvir Khaliq,a Shweta,a Nishi,b
Neena Goyalc and Suman Guptab
aDivision of Medicinal and Process Chemistry, Central Drug Research Institute, Lucknow, India
bDivision of Parasitology, Central Drug Research Institute, Lucknow, India
cDivision of Biochemistry, Central Drug Research Institute, Lucknow, India
Received 24 June 2005; revised 8 July 2005; accepted 8 July 2005
Available online 26 September 2005
Abstract—A large number of novel chromenochalcones were synthesized by pyridine-catalysed chromenylation of mono-chelated
meta-dihydric acetophenones with the monoterpene, citral dimethyl acetal and subsequent Claisen–Schmidt condensation of the
resultant acylchromenes with appropriate aromatic aldehydes. These chromenochalcones 1–19 were screened against in vitro
extracellular promastigotes and intracellular amastigotes of Leishmania donovani. The most potent compound in this series
was compound 9 with a pyridine ring-A, which showed 99% inhibition of promastigotes at 10 lg/ml, 82% at 0.25 lg/ml and
96% at 10 lg/ml concentration against amastigotes.
ꢀ 2005 Elsevier Ltd. All rights reserved.
1. Introduction
non. Licochalcone A (Fig. 1), isolated from Chinese lico-
rice roots, efficiently inhibited the proliferation of L.
donovani and L. major promastigotes and amastigotes
in vitro by inhibiting fumarate reductase,8 a selective tar-
get present in the parasite mitochondria. Recently, we
have described the isolation9 and synthesis10 of few natu-
rally occurring 20,20-dimethyl chromenochalcones and
dihydro chromenochalcones, that is, crotaramosmin,
crotaramin and crotin (Fig. 1) and evaluation of their
antileishmanial activity, which were found to be promis-
ing in our studies. These compounds contain a 2020-di-
Various species of the protozoan parasite Leishmania
cause a broad spectrum of diseases ranging from cutane-
ous healing skin lesions caused by L. major to a fatal
visceral form called kala azar caused by L. donovani.1,2
Leishmaniasis is widespread in many parts of the world
with the highest prevalence in Africa, Asia and Latin
America. It is estimated that 12 million people suffer from
the disease, with 400,000 new cases and 100,000 deaths
each year.3 No fewer than 550 million individuals in 88
countries, including some southern European countries,
are at risk of infection.4 Therapy to treat patients with
leishmaniasis still poses a serious problem. The drugs of
first choice are pentavalent antimonial compounds,
which were developed before 1960,5 and in general,
require long-term treatment and have severe side effects.4
The reported large-scale clinical resistance to antimonial
agents in India and Sudan6 has created an urgent need for
the development of new, efficient and safe drugs for the
treatment of the disease.7 The discovery of new
drugs from traditional medicine is not a new phenome-
R1
OH
O
HO
R2
OCH3
OH
O
O
Licochalcone-A
Crotaramosmin; R1=OH, R2=H
Crotaramin; R1=OCH3, R2=H
Crotin; R1=OH, R2=OH
OCH3
OCH3
O
O
OCH3
OCH3
OH
O
OH
O
Keywords: Chromenochalcones; Antileishmanial activity; Synthesis.
q
2,4-dimethoxy-4'-butoxychalcone (24mbc)
2,4-dimethoxy-4'-allyloxychalcone (24m4ac)
CDRI Comm. No. 6793.
Corresponding author. Tel.: 91 0522 2612411; fax: +91 0522 2623405/
2623938; e-mail: tnarender@rediffmail.com
*
Figure 1. Few naturally occurring and synthetic antileishmanial
chalcones.
0968-0896/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2005.07.005

6544
T. Narender et al. / Bioorg. Med. Chem. 13 (2005) 6543–6550
methyl benzopyran system that is frequently encountered
in many natural products. Some of them have been
reported to exhibit significant biological activities10
including anti-HIV, insecticidal, anti-inflammatory,
etc., and appeared to be more potent antileishmanial
compounds than normal chalcones.10 Several synthetic
chalcones have appeared in the literature, which act as
fumarate reductase inhibitors (FRD)11 and further stud-
ies on antileishmanial chalcones have revealed that large
substitution on the 40-hydroxyl group of ring-B, as in
compounds 2,4-dimethoxy-40-butoxychalcone (24mbc)
and 2,4-dimethoxy-40-allyloxy chalcone (24m4ac)
(Fig. 1), has been shown to be more active.11 We therefore
synthesized a large number of such novel chromenochal-
cones and evaluated their antileishmanial activity.
HO
OH
O
Py
O
O
O
+
O
O
O
1500C; 11h
OMe
OMe
+
OH
OH
OH
6""
23
26
4"
28
27
7""
3""
6"
OCH3
4
1"
3'
2'
5"
4
4'
OCH3
3'"
4'"
2''"
1''"
7"
2"
3"
1'
1
2"
1'"
O
O
3"
2
β
9'
O
O
Aq.KOH/EtOH
1'
β
5'
4"
1"
+
α
α
p-anisaldehyde
8'
3'
6'
OH
10' 6'
OH
25
4'
6"
O
O
7"
17
16
Scheme 2. Synthesis of dicromenochalcone and unusual chalcone.
murine macrophages. These chromenochalcones have
exhibited inhibition of parasites in a concentration rang-
ing from 50 to 0.25 lg/ml. The promastigote inhibition
was 12– 99% at 10 lg/ml, while the inhibition was 34–
99% at 25 lg/ml against amastigotes (Table 1). The most
potent compound in this series was compound 9 with a
pyridine ring-A. It showed 99% inhibition at 10 lg/ml
and 82% at 0.25 lg/ml concentration against promasti-
gotes, whereas the inhibition was 96% at 10 lg/ml against
amastigotes. Compound 1, which has a hydroxyl group at
the 4-position in ring-A, showed significant inhibition of
parasites. The inhibition was 97% at 10 lg/ml and 66%
at 5 lg/ml against promastigotes and 89% at 25 lg/ml
against amastigotes. On the other hand, its dihydroderiv-
ative 19 exhibited diminished activity with 95% at 10 lg/
ml and 31% inhibition at 5 lg/ml in promastigotes and
47% at 25 lg/ml in amastigotes. This revealed that unsat-
uration at a, b-carbons and in the chromene part played a
significant role in enhancing the activity.
2. Synthesis
The general synthesis of chalcones involves the Claisen–
Schmidt condensation of acetophenone with an appro-
priate aldehyde.12 For the preparation of acyl chrom-
enes, the pyridine-catalysed condensation between
citral dimethyl acetal 23 and 2,4-dihydroxyacetophe-
none 22 has been exploited in which one of the two
hydroxyl groups of acetophenone is engaged in chela-
tion (Scheme 1).13 The resultant acyl chromenes 24 were
subjected to Claisen–Schmidt condensation with various
substituted aromatic aldehydes to give chromenochal-
cones (Scheme 1).
To evaluate the activity of dichromenylated chalcones,
we carried out the same chromenylation reaction on
2,4,6-trihydroxy acetophenone 26 and ended up with a
compound 28 with a menthylidene ring system and with
the dichromene derivative 27 (Scheme 2).
The O-alkylated derivatives of compound 1 viz., com-
pounds 2 and 14 did not show any significant activity,
implying the increased role of hydroxyl group in ring-
A as compared to O-alkylated groups. The presence of
nitrogenous substituents either at the 4-position or at
the 3-position in ring-A imparted promising activity to
these chromenochalcones. Compound 3 having a nitro
group at the 4-position exhibited 99% at 10 lg/ml and
86% inhibition at 0.25 lg/ml against promastigotes.
Compound 12 with nitro group at the 3-position also
showed significant results at 10 lg/ml with 99% and
1 lg/ml with 95% inhibition against promastigotes and
at 10 lg/ml with 84% inhibition against amastigotes.
On the contrary, the presence of halogens in ring-A
did not impart any significant activity. Compound 4
showed 86% at 10 lg/ml and no inhibition at all below
this concentration in promastigotes. There was no inhi-
bition against amastigotes, even at 50 lg/ml concentra-
tion. These findings reveal that substituent effect in
ring-A is pivotal in determining the antileishmanial
activity of chromenochalcones.
3. In vitro antileishmanial activity
Chalcones, natural14 as well as synthetic, have been
reported as potential antileishmanial agents.15 Based on
ourearlierfindings,10 we were promptedtosynthesizevar-
ious substituted chromenochalcones 1–19 (Table 1) to im-
prove the antileishmanial activity. All the compounds
were tested against extracellular promastigotes16 and
intracellular amastigotes17 of L. donavani residing within
O
HO
Py
+
1500 C; 7h
OMe
OMe
OH
24
O
OH
O
23
22
6"
7"
OCH3
Aq.KOH/EtOH
OHC
4
OCH3
OCH3
25
4"
Ring-B in these chromenochalcones is also contributing
to antileishmanial activity against the parasites. This
was revealed by compound 18 in which the chromene
group was absent, showing 60 and 15% inhibition at
50 lg/ml against promastigotes and amastigotes, respec-
tively. Whereas its chromenylated derivative 2 showed
87 and 49% inhibition at the same concentration. Thus,
1'
8'
O
2"
1"
O
7'
β
Pd/C/H2
MeOH/6h
1
3'
α
5'
OH
4'
OH
O
O
20
2
Scheme 1. Synthesis of chromenochalcones and chromanodihydro-
chalcones.

T. Narender et al. / Bioorg. Med. Chem. 13 (2005) 6543–6550
Table 1. In vitro antileishmanial activity of synthetic chromenochalcones
6545
O
A
B
Compound
A
B
Concentration
(lg/ml)
Promastigotes
(%inhibition)
Concentration
(lg/ml)
Amastigotes
(%inhibition)
O
O
O
O
O
O
O
O
O
O
O
OH
1
10
50
10
10
10
10
10
10
10
10
10
10
10
10
97
87
99
86
95
80
5
25
50
25
25
25
25
25
25
25
25
25
25
25
25
89
OH
OH
OH
OH
OH
OH
OCH
3
2
3
49
34
NO
Cl
F
2
4
NIa
NI
NI
NI
NI
99
5
Br
6
N
Cl
7
OH
OH
OH
OH
OH
8
20
99
85
91
99
49
16
Cl
N
9
Cl
10
11
12
13
14
NI
NI
93
Cl
OMe
OMe
OMe
O
NO2
OH
O
NI
NI
OCH3
OH
O
OCH2CH3
OH
(continued on next page)

6546
T. Narender et al. / Bioorg. Med. Chem. 13 (2005) 6543–6550
Table 1 (continued)
Compound
A
B
Concentration
(lg/ml)
Promastigotes Concentration Amastigotes
(%inhibition)
(%inhibition) (lg/ml)
O
O
15
10
10
10
99
12
24
25
25
25
NI
NO
OH
OH
O
OCH3
16
17
NI
31
O
O
OCH
3
OH
O
HO
OMe
18
19
20
21
50
10
60
98
90
45
50
25
50
50
15
47
92
21
OH
OH
O
OH
O
H2N
HN
NH2
NH
O
O
1
Pentamidine
CH OH
2
CHOH
CH OH
2
CHOH
-
OH
Sb Sb
O
H
H
H
O
O
O
O
O
O
H
H
H
3Na+ 9H O
2
500
-
-
COO
COO
Sodium stibogluconate
a NI, no inhibition.
introduction of a chromene ring with its 5-membered side
chain at the 40position in ring-B led to enhanced activity.
NMR spectra were recorded on a Bruker WM
200 MHz spectrometer in deuterated chloroform with
TMS as internal reference. IR spectra of the com-
pounds were recorded on a Perkin–Elmer AC-1 spec-
trometer using KBr pellets. FAB mass spectra of all
compounds were measured with a Jeol SX 102/DA
6000 mass spectrometer using a beam of Argon (2–
8 ev). Microanalysis were determined on a Carlo Erba
EA-1108 element analyzer. Thin-layer chromatography
was performed on Merck pre-coated silica gel plates.
For column chromatography, silica gel of 60–120
mesh was used.
Dichromenochalcones viz., compounds 16 and 17 did
not exhibit any significant activity, which suggests that
monochromene ring is vital in determining the
antileishmanial potential of these chromenochalcones.
4. Conclusion
We have synthesized few naturally occurring chalcones,
chromenochalcones and synthetic chromenochalcones
and investigated their antileishmanial activity to provide
a scientific rationale for the antiprotozoal potency of
plants used in ethnomedicine in the search of new antipro-
tozoaldrugs. Ourresultsdisclosethatchromenochalcones
have potent antileishmanial activity and further work is in
progress to improve the potency of these compounds.
5.2. General procedure for the synthesis of 1-(5-hydroxy-
2-methyl-2-(4-methylpent-3-enyl)-2H-chromen-6-yl)
ethanone (24)
To a stirred solution of 2,4-dihydroxyacetophenone 22
(10.64 g, 0.07 mol) in pyridine (9.48 g, 0.12 mol) was
added citral dimethyl acetal 23 (13.66 g, 0.035 mol)
slowly. The mixture was refluxed for 4 h at 150 ꢁC.
More citral dimethyl acetal (13.66 g, 0.035 mol) was
added and refluxed for further 3 h. The reaction mix-
ture was concentrated under reduced pressure and the
resultant concentrate was subjected to silica gel chro-
matography using hexane/ethyl acetate (94:6) system
as eluent.
5. Experimental
5.1. General
Melting points were recorded on a Buchi-530 capillary
melting point apparatus and are uncorrected. 1H

T. Narender et al. / Bioorg. Med. Chem. 13 (2005) 6543–6550
6547
5.3. General procedure for the synthesis of dichromene of
2,4,6-trihydroxyacetophenone (27)
CDCl3) d 1.42 (s, 3H, H-100), 1.56 (s, 3H, H-600), 1.67 (s,
3H, H-700), 1.72 (t, J = 6 Hz, 2H, H-200), 2.08 (q,
J = 6 Hz, 2H, H-300), 3.83 (s, 3H, OMe), 5.08 (t,
J = 6 Hz, 1H, H-400), 5.52 (d, J = 10.2 Hz, 1H, H-30),
6.35 (d, J = 8.8 Hz, 1H, H-70), 6.78 (d, J = 10.1 Hz, 1H,
H-40), 6.90 (d, J = 8.4 Hz, 2H, H-3,5), 7.41 (d,
J = 15.4 Hz, 1H, H-a), 7.58 (d, J = 8.4 Hz, 2H, H-2,6),
7.69 (d, J = 8.8 Hz, 1H, H-80), 7.83 (d, J = 15.4 Hz, 1H,
H-b). Anal. Calcd for C26H28O4: C, 77.20; H, 6.98; O,
15.82. Found: C, 77.40; H, 7.10.
To a stirred solution of 2,4,6-trihydroxyacetophenone 26
(3.36 g, 0.02 mol) in pyridine (4.74 g, 0.06 mol) was added
citral dimethyl acetal 23 (3.96 g, 0.02 mol) slowly. The
mixture was refluxed for 6 h at 150 ꢁC. More citral
dimethyl acetal (1.98 g, 0.01 mol) was added and refluxed
for further 5 h. The reaction mixture was concentrated
under reduced pressure and the resultant concentrate
was subjected to silica gel chromatography using hex-
ane/ethyl acetate (96:4) system as eluent.
5.8. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)- 3-(4-nitro-phenyl)-propenone (3)
5.4. General procedure for the synthesis of chromenoch-
alcones (2)
Yield: 30%; mp 140 ꢁC; FAB MS 420 (M+1); IR (KBr)
3425 cmꢀ1 (OH), 1638 cmꢀ1 (CO); 1H NMR (200 MHz,
CDCl3) d 1.44 (s, 3H, H-100), 1.56, (s, 3H, H-600), 1.65 (s,
3H, H-700), 1.73 (t, J = 4 Hz, 2H, H-200), 2.08 (q,
J = 8 Hz, 2H, H-300), 5.09 (t, J = 6 Hz, 1H, H-400), 5.54
(d, J = 10.1 Hz, 1H, H-30), 6.36 (d, J = 8.8 Hz, 1H,
H-70), 6.77 (d, J = 10.1 Hz, 1H, H-40), 7.63 (d,
J = 15.5 Hz, 1H, H-a), 7.69 (d, J = 8.7 Hz, 2H, H-2,6),
7.76 (d, J = 8.8 Hz, 1H, H-80), 7.85 (d, J = 15.5 Hz,
1H, H-b), 8.26 (d, J = 8.7 Hz, 2H, H-3,5). Anal. Calcd
for C25H25NO5: C, 77.58; H, 6.01; N, 3.34; O, 19.07.
Found: C, 77.62; H, 6.07; N, 3.39.
To a solution of chromeno/dichromeno acetophenone
24:28 (0.500 g, 0.02 mol) in aqueous potassium hydrox-
ide (2-pellets) in ethanol (5 ml) was added the aldehyde
25 (0.04 mol). The mixture was kept at room tempera-
ture for 48 h. The resultant mixture was quenched in
ice-cold water, acidified with 1 N HCl. The crude prod-
uct was filtered under suction and extracted with ethyl
acetate and organic layer concentrated under reduced
pressure. The resultant concentrate was subjected to sil-
ica gel column chromatography using different polarity
systems of hexane/ethyl acetate.
5.9. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(4-chloro-phenyl)-propenone (4)
5.5. General procedure for the synthesis of nitro-
chromenochalcones (3, 12)
Yield: 38%; mp 130 ꢁC ; FAB MS 409 (M+1); IR (KBr)
3435 cmꢀ1 (OH), 1640 cmꢀ1 (CO); 1H NMR (200 MHz,
CDCl3) d 1.35 (s, 3H, H-100), 1.49 (s, 3H, H-600), 1.65 (s,
3H, H-700), 1.72 (t, J = 6 Hz, 2H, H-200), 2.04 (q,
J = 6 Hz, 2H, H-300), 5.01 (t, J = 6 Hz, 1H, H-400), 5.45
(d, J = 10.1 Hz, 1H, H-30), 6.29 (d, J = 8.7 Hz, 1H,
H-70), 6.70 (d, J = 10.1 Hz, 1H, H-40), 7.30 (d,
J = 8.2 Hz, 2H, H-3,5), 7.46 (d, J = 15.4 Hz, 1H, H-a),
7.48 (d, J = 8.2 Hz, 2H, H-2,6), 7.60 (d, J = 8.8 Hz,
1H, H-80), 7.72 (d, J = 15.4 Hz, 1H, H-b). Anal. Calcd
for C25H25O3Cl: C, 73.43; H, 6.16; O, 11.74; Cl, 8.67.
Found: C, 73.31; H, 6.18.
To a stirred solution of chromene of 2,4-dihydroxyace-
tophenone 25 (0.572 g, 0.002 mol) in THF (5 ml) was
added sodium hydride (0.115 g, 0.005 mol) slowly and
stirred at rt for 20 min. Then the nitro substituted alde-
hyde (0.302 g, 0.002 mol) in THF was added and the
mixture was stirred at rt under nitrogen for 8 h. The
mixture was poured into ice-cold water, acidified with
1 N HCl. Filtration, extraction with ethyl acetate and
concentration under reduced pressure and silica gel col-
umn chromatography led to isolation of the desired
compound. (eluent: hexane/ethyl acetate).
5.6. 1-(5-Hydroxy–2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(4-hydroxy-phenyl)-propenone (1)
5.10. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(4-fluoro-phenyl)-propenone (5)
Yield: 44%; semisolid; FAB MS 391 (M+1); IR (KBr)
Yield: 37%; mp 110 ꢁC; FAB MS 392 (M+1); IR (KBr)
3424 cmꢀ1 (OH), 1640 cmꢀ1 (CO); 1H NMR (200 MHz,
CDCl3) d 1.43 (s, 3H, H-100), 1.56 (s, 3H, H-600), 1.65 (s,
3H, H-700), 1.73 (t, J = 4 Hz, 2H, H-200), 2.10 (q,
J = 8 Hz, 2H, H-300), 5.09 (t, J = 6 Hz, 1H, H-400), 5.54
(d, J = 10.1 Hz, 1H, H-30), 6.37 (d, J = 8.8 Hz, 1H,
H-70), 6.79 (d, J = 10.1 Hz, 1H, H-40), 7.12 (t,
J = 8.4 Hz, 2H, H-3,5), 7.46 (d, J = 15.4 Hz, 1H, H-a),
7.61 (d, J = 8.4 Hz, 2H, H-2,6), 7.63 (d, J = 8.8 Hz,
1H, H-80), 7.83 (d, J = 15.4 Hz, 1H, H-b). Anal. Calcd
for C25H25O3F.
1
3436 cmꢀ1 (OH), 1635 cmꢀ1 (CO); H NMR(200 MHz,
CDCl3) d 1.43 (s, 3H, H-100), 1.56 (s, 3H, H-600), 1.65 (s,
3H, H-700), 1.73 (t, J = 4 Hz, 2H, H-200), 2.08 (q,
J = 8 Hz, 2H, H-300), 5.09 (t, J = 8 Hz, 1H, H-400), 5.53
(d, J = 10.1 Hz, 1H, H-30), 6.37 (d, J = 8.8 Hz, 1H, H-
70), 6.79 (d, J = 10.1 Hz, 1H, H-40), 6.88 (d, J = 8.4 Hz,
2H, H-3,5), 7.42 (d, J = 15.3 Hz, 1H, H-a), 7.53 (d,
J = 8.4 Hz, 2H, H-2,6), 7.70 (d, J = 8.8 Hz, 1H, H-80),
7.82 (d, J = 15.3 Hz, 1H, H-b). Anal. Calcd for C25H26
O4: C, 76.90; H, 6.71; O, 16.39. Found: C, 76.80; H, 6.69.
5.7. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(4-methoxy-phenyl)-propenone (2)
5.11. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(4-bromo-phenyl)-propenone (6)
Yield: 50%; semisolid; FAB MS 405 (M+1); IR (KBr)
3435 cmꢀ1 (OH), 1635 cmꢀ1 (CO); 1H NMR (200 MHz,
Yield: 46%; mp 96 ꢁC; FAB MS 454 (M+1); IR (KBr)
3428 cmꢀ1 (OH), 1635 cmꢀ1 (CO); 1H NMR (200 MHz,

6548
T. Narender et al. / Bioorg. Med. Chem. 13 (2005) 6543–6550
CDCl3) d 1.43 (s, 3H, H-100), 1.56 (s, 3H, H-600), 1.65 (s, 3H,
H-700), 1.73 (t, J = 4Hz, 2H, H-200), 2.08 (q, J = 8Hz, 2H,
H-300), 5.08 (t, J = 6 Hz, 1H, H-400), 5.54 (d, J = 10.1 Hz,
1H, H-30), 6.36 (d, J = 8.8 Hz, 1H, H-70), 6.77 (d,
J = 10.1 Hz, 1H, H-40), 7.48 (d, J = 8.8 Hz, 2H, H-2,6),
7.50 (d, J = 8.8 Hz, 2H, H-3,5), 7.52 (d, J = 15.4 Hz,
1H, H-a), 7.68 (d, J = 8.8 Hz, 1H, H-80), 7.78 (d,
J = 15.4 Hz, 1H, H-b). Anal. Calcd for C25H25O3Br: C,
66.23; H, 5.56; Br, 17.62; O, 10.59. Found: C, 65.98; H,
5.23.
5.15. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(3,5-dichloro-phenyl)-propenone (10)
Yield: 26%; mp 92–95 ꢁC; FAB MS 443 (M+1); IR
(KBr) 3429 cmꢀ1 (OH), 1640 cmꢀ1 (CO); 1H NMR
(200 MHz, CDCl3) d 1.43 (s, 3H, H-100), 1.56 (s, 3H,
H-600), 1.65 (s, 3H, H-700), 1.73 (t, J = 4 Hz, 2H, H-200),
2.08 (q, J = 8 Hz, 2H, H-300), 5.07 (t, J = 6 Hz, 1H,
H-400), 5.53 (d, J = 10.1 Hz, 1H, H-30), 6.37 (d,
J = 8.8 Hz, 1H, H-70), 6.77 (d, J = 10.1 Hz, 1H, H-40),
7.41 (d, J = 1.6 Hz, 1H, H-6), 7.44 (d, J = 1.6 Hz, H-
2), 7.50 (d, J = 15.0 Hz, 1H, H-a), 7.68 (d, J = 8.3 Hz,
1H, H-80), 7.74 (d, J = 15.0 Hz, 1H, H-b). Anal. Calcd
for C25H24O3Cl2: C, 67.73; H, 5.46; Cl, 15.99; O,
10.83. Found: C, 67.64; H, 5.20.
5.12. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(4,4,N,N-dimethylamino-phenyl)-prope-
none (7)
Yield: 48%; semisolid; FAB MS 418 (M+1); IR (KBr)
3435 cmꢀ1 (OH), 1635 cmꢀ1 (CO); 1H NMR
(200 MHz, CDCl3) d 1.35 (s, 3H, H-100), 1.49 (s, 3H,
H-600), 1.51 (s, 3H, H-700), 1.50 (t, J = 6 Hz, 2H, H-200),
2.04 (q, J = 6 Hz, 2H, H-300), 2.96 (s, 6H, NMe2), 5.08
(t, J = 6 Hz, 1H, H-400), 5.44 (d, J = 10.1 Hz, 1H,
H-30), 6.27 (d, J = 8.8 Hz, 1H, H-70), 6.60 (d,
J = 8.4 Hz, 2H, H-3,5), 6.72 (d, J = 10.1 Hz, 1H,
H-40), 7.27 (d, J = 15.6 Hz, 1H, H-a), 7.47 (d,
J = 8.4 Hz, 2H, H-2,6), 7.63 (d, J = 8.8 Hz, 1H, H-80),
7.78 (d, J = 15.6 Hz, 1H, H-b). Anal. Calcd for C27
H31NO3: C, 77.67; H, 7.48; N, 3.35; O, 11.50. Found:
C, 77.50; H, 7.27; N, 3.15.
5.16. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(3,4,5-trimethoxy-phenyl)-propenone
(11)
Yield: 30%; semisolid; FAB MS 465 (M+1); IR (KBr)
3424 cmꢀ1 (OH), 1638 cmꢀ1 (CO); 1H NMR (200 MHz,
CDCl3) d 1.43 (s, 3H, H-100), 1.57 (s, 3H, H-600), 1.65 (s,
3H, H-700), 1.73 (t, J = 4 Hz, 2H, H-200), 2.12 (q,
J = 6 Hz, 2H, H-300), 3.91 (s, 9H, OMe), 5.08 (t,
J = 6 Hz, 1H, H-400), 5.54 (d, J = 10.1 Hz, 1H,
H-30), 6.38 (d, J = 8.8 Hz, 1H, H-70), 6.80 (d,
J = 10.1 Hz, 1H, H-40), 6.86 (s, 1H, H-2), 7.26 (s, 1H, H-
6), 7.41 (d, J = 15.3 Hz, 1H, H-a), 7.72 (d, J = 8.8 Hz,
1H, H-80), 7.79 (d, J = 15.3 Hz, 1H, H-b). Anal. Calcd
for C28H32O6: C, 72.39; H, 6.94; O, 20.66. Found: C,
72.40; H, 6.98.
5.13. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(2,4-dichloro-phenyl)-propenone (8)
Yield: 26%; mp 76 ꢁC; FAB MS 443 (M+1); IR (KBr)
3433 cmꢀ1 (OH), 1640 cmꢀ1 (CO); 1H NMR
(200 MHz, CDCl3) d 1.41 (s, 3H, H-100), 1.56 (s, 3H,
H-600), 1.68 (s, 3H, H-700), 1.73 (t, J = 6Hz, 2H, H-200),
2.08 (q, J = 6 Hz, 2H, H-300), 5.07 (t, J = 6 Hz, 1H,
H-400), 5.54 (d, J = 10.1 Hz, 1H, H-30), 6.36 (d,
J = 8.9 Hz, 1H, H-70), 6.79 (d, J = 10.1 Hz, 1H, H-40),
7.29 (dd, J = 8.2, 1.8 Hz, 1H, H-5), 7.46 (d, J = 1.8 Hz,
1H, H-3), 7.50 (d, J = 15.4 Hz, 1H, H-a), 7.63 (d,
J = 8.9 Hz, 1H, H-80), 7.68 (d, J = 8.2 Hz, 1H, H-6),
8.16 (d, J = 15.4 Hz, 1H, H-b). Anal. Calcd for
C25H24O3Cl2: C, 67.73; H, 5.46; Cl, 15.99; O, 10.83.
Found: C, 67.40; H, 5.18.
5.17. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(3-nitro-phenyl)-propenone (12)
Yield: 30%; mp: 115–118 ꢁC MS 420 (M+1); IR (KBr)
3430 cmꢀ1 (OH), 1640 cmꢀ1 (CO); 1H NMR (200 MHz,
CDCl3) d 1.36 (s, 3H, H-100), 1.49 (s, 3H, H-600), 1.58 (s,
3H, H-700), 1.68 (t, J = 4 Hz, 2H, H-200), 2.03 (q,
J = 8 Hz, 2H, H-300), 5.01 (t, J = 8 Hz, 1H, H-400), 5.47
(d, J = 10.1 Hz, 1H, H-30), 6.33 (d, J = 8.9 Hz, 1H, H-
70), 6.70 (d, J = 10.1 Hz, 1H, H-40), 7.58 (d, J = 15.4 Hz,
1H, H-a), 7.60 (d, J = 8.9 Hz, 1H, H-80), 7.64 (dd,
J = 5.9 Hz, 1H, H-5), 7.80 (d, J = 15.4 Hz, 1H, H-b),
8.18 (dd, J = 9.9, 1.9 Hz, 1H, H-6) (d, J = 8.1 Hz, 1H,
H-4), 8.43 (s, 1H, H-2). Anal. Calcd for C25H25NO5: C,
77.58; H, 6.01; N, 3.34; O, 19.07. Found: C, 77.64; H,
6.05; N, 3.39.
5.14. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(3-pridyl)-propenone (9)
Yield: 46%; semisolid; FAB MS m/z 376 (M+1); IR
(KBr) 3425 cmꢀ1 (OH), 1638 cmꢀ1 (CO); 1H NMR
(200 MHz, CDCl3) d 1.05 (s, 3H, H-100), 1.15 (s, 3H,
H-600), 1.36 (s, 3H, H-700), 1.49 (t, J = 5 Hz, 2H,
H-200), 2.05 (q, J = 8 Hz, 2H, H-300), 5.01 (t,
J = 8 Hz, 1H, H-400), 5.46 (d, J = 10.1 Hz, 1H, H-30),
6.31 (d, J = 8.8 Hz, 1H, H-70), 6.71 (d, J = 10.1 Hz,
1H, H-40), 7.29 (dd, J = 7.7 Hz, 1H, H-5), 7.54 (d,
J = 15.6 Hz, 1H, H-a), 7.61 (d, J = 8.8 Hz, 1H,
H-80), 7.77 (d, J = 15.5 Hz, 1H, H-b), 7.87 (d,
J = 7.4 Hz, 1H, H-6), 8.54 (d, J = 3.4 Hz, 1H, H-4),
8.79 (s, 1H, H-2). Anal. Calcd for C24H25NO3: C,
76.77; H, 6.71; N, 3.73; O, 12.78. Found: C, 76.82;
H, 6.76; N, 3.79.
5.18. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(2-methoxy-phenyl)-propenone (13)
Yield: 41%; semisolid; FAB MS 405 (M+1); IR (KBr)
3428 cmꢀ1 (OH), 1640 cmꢀ1 (CO); 1H NMR
(200 MHz, CDCl3) 1.43 (s, 3H, H-100), 1.57 (s, 3H,
H-600), 1.65 (s, 3H, H-700), 1.73 (t, J = 4 Hz, 2H, H-200),
2.12 (q, J = 8 Hz, 2H, H-300), 3.92 (s, 3H, OMe), 5.08
(t, J = 8 Hz, 1H, H-400), 5.53 (d, J = 10.1 Hz, 1H, H-
30), 6.36 (d, J = 8.8 Hz, 1H, H-70), 6.80 (d,
J = 10.1 Hz, 1H, H-40), 6.97 (dd, J = 8.6 Hz, 2H, H-
2,6), 7.37 (d, J = 15.5 Hz, 1H, H-a), 7.64 (dd, J = 6,
10 Hz, 2H, H-3,5), 7.70 (d, J = 8.6 Hz, 1H, H-80), 8.18

T. Narender et al. / Bioorg. Med. Chem. 13 (2005) 6543–6550
6549
(d, J = 15.5 Hz, 1H, H-b). Anal. Calcd for C26H28O4: C,
77.20; H, 6.98; O, 15.82. Found: C, 77.30; H, 7.05.
J = 8.8 Hz, 2H, H-2,6), 7.75 (d, J = 15.6 Hz, 1H, H-a),
8.14 (d, J = 15.6 Hz, 1H, H-b).
5.19. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(4-ethoxy-phenyl)-propenone (14)
5.23. 1-(2,4-dihydroxy-phenyl)-3-(4-methoxy-phenyl)-prope-
none (18)
Yield: 29%; semisolid; FAB MS 419 (M+1); IR (KBr)
3434 cmꢀ1 (OH), 1636 cmꢀ1 (CO); 1H NMR
(200 MHz, CDCl3) d 1.42 (s, 3H, H-100), 1.56 (s, 3H,
H-600), 1.67 (s, 3H, H-700), 1.71 (t, J = 8 Hz, 3H,
OCH2CH3), 1.77 (t, J = 4 Hz, 2H, H-200), 2.08 (q,
J = 8 Hz, 2H, H-300), 4.05 (q, J = 8 Hz, 2H, OCH2CH3),
5.08 (t, J = 6 Hz, 1H, H-400), 5.52 (d, J = 10.1 Hz, 1H, H-
30), 6.35 (d, J = 8.8 Hz, 1H, H-70), 6.78 (d, J = 10.1 Hz,
1H, H-40), 6.89 (d, J = 8.6 Hz, 2H, H-3,5), 7.40 (d,
J = 14.9 Hz, 1H, H-a), 7.56 (d, J = 8.8 Hz, 2H, H-2,6),
7.69 (d, J = 8.6 Hz, 1H, H-80), 7.83 (d, J = 14.9 Hz,
1H, H-b). Anal. Calcd for C27H30O4: C, 77.48; H,
7.22; O, 15.29. Found: C, 77.2; H, 7.02.
Yield: 14%; FAB MS 271 (M+1); IR (KBr) 3436 cmꢀ1
1
(OH), 1640 cmꢀ1 (CO); H NMR (200 MHz, CDCl3) d
3.87 (s, OMe, 3H), 6.27 (d, J = 2Hz, 1H, H-30), 6.39
(dd, J = 8.6, 2 Hz, 1H, H-50), 6.83 (d, J = 8.4 Hz, 2H,
H-3,5), 7.41 (d, J = 15.4 Hz, 1H, H-a), 7.62 (d,
J = 8.4 Hz, 2H, H-2,6), 7.73 (d, J = 15.4 Hz, 1H, H-b),
8.14 (d, J = 8.6 Hz, 1H, H-60). Anal. Calcd for
C16H14O4: C, 71.10; H, 5.22; O, 23.68.
5.24. 1-(5-Hydroxy-2-methyl-2-(4-methylpentanyl)2H-
chroman-6-yl)-3-(4-hydroxy-phenyl)-propenone (19)
Yield: 73%; semisolid; FAB MS 397 (M+1); IR (KBr)
3436 cmꢀ1 (OH), 1650 cmꢀ1 (CO); 1H NMR
(200 MHz, CDCl3) d 0.77 (s, 3H, H-600), 0.88 (s, 3H,
H-700), 1.17 (m, 2H, H-300), 1.19 (m, 2H, H-400), 1.21 (s,
3H, H-100), 1.43–152 (m, 3H, H-200,500), 1.72 (t,
J = 6.02 Hz, 2H, H-30), 2.58 (t, J = 6.3 Hz, 2H, H-40),
2.90 (t, J = 6.8 Hz, 2H, H-b), 3.07 (t, J = 8 Hz, 2H,
H-a), 6.23 (d, J = 10 Hz, 1H, H-80), 6.68 (d,
J = 8.2 Hz, 2H, H-3,5), 7.00 (d, J = 8.1 Hz, 2H, H-2,6),
7.41 (d, J = 8.5 Hz, 1H, H-70). Anal. Calcd for
C25H32O4: C, 75.73; H, 8.13; O, 16.14. Found: C,
75.80; H, 8.20.
5.20. 1-(5-Hydroxy-2-methyl-2-(4-methylpent-3-enyl)2H-
chromen-6-yl)-3-(4-cyano-phenyl)-propenone (15)
Yield: 29%; mp 130 ꢁC; FAB MS 400 (M+1); IR (KBr)
3435 cmꢀ1 (OH), 1635 cmꢀ1 (CO); 1H NMR (200 MHz,
CDCl3) d 1.43 (s, 3H, H-100), 1.56 (s, 3H, H-600), 1.65 (s,
3H, H-700), 1.73 (t, J = 4 Hz, 2H, H-200), 2.08 (q,
J = 8 Hz, 2H, H-300), 5.08 (t, J = 6 Hz, 1H, H-400), 5.55
(d, J = 10.1 Hz, 1H, H-30), 6.38 (d, J = 8.8 Hz, 1H, H-
70), 6.77 (d, J = 10.1 Hz, 1H, H-40), 7.61 (d,
J = 15.4 Hz, 1H, H-a), 7.65 (d, J = 7.6 Hz, 2H, H-3,
5), 7.67 (d, J = 7.6 Hz, 2H, H-2,6), 7.82 (d, J = 15.4,
1H, H-b) Anal. Calcd for C26H25NO3: C, 78.17; H,
6.31; N, 3.51; O, 12.02. Found: C, 78.12; H, 6.17; N,
3.39.
5.25. Procedure for in vitro antileishmanial screening
5.25.1. Antipromastigote activity. Luciferase transfected
L. donovani promastigotes (MHOM/IN/80/Dd-8, ob-
tained from Imperial College, London), which are more
stable under the influence of G 418,16 were maintained
at 25 1 ꢁC in medium 199 (Sigma Chemical, USA)
supplemented with 10% foetal calf serum (Gibco). The
in vitro effect of compounds on the growth of promasti-
gotes was assessed by monitoring the luciferase activity
of viable cells after treatment. The transgenic promasti-
gotes of late log phase were seeded at 5 · 105/100 ll
medium 199/well in 96-well flat-bottomed microtitre
(MT) plates (CELLSTAR) and incubated for 72 h in
medium alone or in the presence of serial dilutions of
drugs (0.25–10 lg/ml) in DMSO.16 Parallel dilutions of
DMSO were used as controls. After incubation, an ali-
quot (50 ll) of promastigote suspension was aspirated
from each well of a 96-well plate and mixed with an
equal volume of Steady Glo(R) reagent (Promega) and
luminescence was measured by a luminometer. The val-
ues were expressed as relative luminescence unit (RLU).
5.21. 1-(5-Hydroxy-2,8-dimethyl-2,8-bis-(4-methylpent-3-
enyl)-2H,8H-pyrano(2,3-f)chromen-6-yl)-3-(4-methoxy-
phenyl)-propenone (16)
Yield: 51%; semisolid; FAB MS; 555 (M+1); IR (KBr)
3435 cmꢀ1 (OH), 1636 cmꢀ1 (CO); 1H NMR
(200 MHz, CDCl3) d 1.42 (s, 3H, H-100), 1.49 (s, 6H,
H-60060000), 1.57 (s, 3H, H-10000), 1.64 (s, 6H, H-700,70000),
1.83 (t, J = 4 Hz, 4H, H-200,20000), 2.08 (q, J = 8 Hz, 4H,
300,30000), 3.85 (s, 3H, OMe), 5.09 (t, J = 6 Hz, 2H,
H-400,40000), 5.40 (d, J = 10.1 Hz, 1H, H-30), 5.44 (d,
J = 10.1 Hz, 1H, H-3000), 6.65 (d, J = 10.1Hz, 1H,
H-40), 6.73 (d, J = 10.1 Hz, 1H, H-4000), 6.91 (d,
J = 8.8 Hz, 2H, H-3,5), 7.54 (d, J = 8.8 Hz, 2H, H-2,6),
7.75 (d, J = 15.6 Hz, 1H, H-a), 7.98 (d, J = 15.6 Hz,
1H, H-b). Anal. Calcd for C36H42O5: C, 77.95; H,
7.63; O, 14.42. Found: C, 77.90; H, 7.58.
5.22. Methylidene ring derivative (17)
Percentage inhibition ¼ N-n=NX100;
where N is average relative luminescence unit (RLU) of
control wells; n is average RLU of treated wells.
Yield: 53%; mp 190–192 ꢁC; FAB MS m/z 421 (M+1);
1
IR (KBr) 3435 cmꢀ1 (OH), 1635 cmꢀ1 (CO); H NMR
(200 MHz, CDCl3) d 1.06 (s, 3H, H-600), 1.25–1.32 (m,
2H, H-300), 1.39 (s, 3H, H-700), 1.44 (m, 1H, H-500), 1.65
(s, 3H, H-100), 1.89–1.90 (m, 2H, H-200), 2.12–2.17 (m,
2H, H-30), 2.77 (t, 1H, H-50), 3.84 (s, 3H, OMe), 6.09
(s, 1H, ArH), 6.93 (d, J = 8.8 Hz, 2H, H-3,5), 7.57 (d,
5.25.2. Antiamastigotes activity. For assessing the activity
of compounds against the amastigote stage of the para-
site, mouse macrophage cell line (J-774A.1) infected with
promastigotes expressing luciferase firefly reporter gene

6550
T. Narender et al. / Bioorg. Med. Chem. 13 (2005) 6543–6550
was used.17 Cells were seeded in a 96-well plate
(5 · 104 cells/100 ll/well) in RPMI-1640 containing 10%
foetal calf serum and the plates were incubated at 37 ꢁC
in a CO2 incubator. After 24 h, the medium was replaced
with fresh medium containing stationary-phase promasti-
gotes (2.5 · 105/100 ll/well). Promastigotes invade the
macrophage and are transformed into amastigotes. The
test material in appropriate concentrations (50 and
10 lg/ml) in complete medium was added after replacing
the previous medium and the plates were incubated at
37 ꢁC in a CO2 incubator for 24 h or more. After incuba-
tion, the drug-containing medium was decanted and 50 ll
PBS was added in each well and mixed with an equal vol-
ume of steady Glo reagent. After gentle shaking for 1–
2 min, the reading was taken in a luminometer.
4. Olliaro, P. L.; Bryceson, A. D. M. Parasitol. Today 1993,
9, 323.
5. Neal, R. A., Peters, W., Kendrick, K., Eds. The Leish-
maniasis in Biology and Medicine; Academic Press:
London, 1987; p 793.
6. United Nations Development Program World Health
Organisation, Special Programme for Research and
Training in Tropical Diseases (TDR). TDR News 1990,
34, 7.
7. Megregor, A. Lancet 1998, 351, 575.
8. Chen, M.; Christensen, S. B.; Blom, J.; Lemmich, E.;
Nadelmann, L.; Fich, K.; Theander, T. G.; Kharazmi, A.
Antimicrob. Agents Chemother. 1993, 37, 2550.
9. Kumar, J. K.; Narender, T.; Rao, M. S.; Rao, P. S.; Toth,
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2023.
Acknowledgments
The authors are grateful to the Director, CDRI, Luc-
know, for constant encouragement for the programme
on antileishmanial drugs. We also thank the RSIC for
spectral data and J. P. Chaturvedi for technical support.
Tanvir and Shweta are thankful to ICAR, and DOD,
New Delhi, respectively, for financial support.
12. Wattanasin, S.; Murphy, W. S. Synthesis 1980, 647.
13. Bandaranayake, W. M.; Crombie, O.; Whiting, D. A.
J. Chem. Soc. (C) 1971, 804.
14. Kayser, O.; Kiderlen, A. F. Phytother. Res. 2001, 15,
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15. Nielsen, S. F.; Christensen, S. B.; Cruciani, G.; Kharaz-
ami, A.; Liljefors, T. J. Med. Chem. 1998, 41, 4819.
16. Croft, S. L.; Snowdon, D.; Yardly, V. J. Antimicrob.
Agents Chemother. 1996, 38, 1041.
17. Gupta, S.; Ashitosh; Ramesh; Goyal, N. Development of
rapid, sensitive semi in vivo model using Leishmania
donovani cell lines expressing luciferase reporter gene,
Abstract presented in 9th National Conference on Bioac-
tive Hyterocycles and Drug Discovery Paradigm, held at
Saurastra University, Department of Chemistry, Rajkot,
Gujarat, India, Jan 8–10, 2005.
References and notes
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