A rapid access to new coumarinyl chalcone and substituted chromeno[4,3-c]pyrazol-4(1H)-ones and their antibacterial and DPPH radical scavenging activities

Article abstract of DOI:10.1007/s00044-010-9326-1

A series of new coumarin derivatives 4 containing a chalcone moiety were synthesized by condensation of 3-acetyl-4-hydroxycoumarin 1 with aryl or heteroaryl aldehydes 2 in the presence of piperidine in chloroform. The interaction of 3-formyl-4-chloro-coumarin 3 with nitrogen compound nucleophiles are described and lead to the corresponding substituted chromen[4,3-c] pyrazol-4-ones 5. The structures of the obtained compounds were established on the basis of IR|1D|2D NMR, while crystal structure of 3-acetyl-4-hydroxy coumarin 1 was determined using X-ray diffraction and further were evaluated for possible antibacterial and antioxidant activities. The coumarinic chalcone 4d has been found to be the most active (IC₅₀ = 2.07 μM) in this study.

Full text of DOI:10.1007/s00044-010-9326-1

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2011) 20:522–530
DOI 10.1007/s00044-010-9326-1
REVIEW
A rapid access to new coumarinyl chalcone and substituted
chromeno[4,3-c]pyrazol-4(1H)-ones and their antibacterial
and DPPH radical scavenging activities
•
Naceur Hamdi Cedric Fischmeister
•
´
M. Carmen Puerta Pedro Valerga
•
Received: 14 July 2009 / Accepted: 12 February 2010 / Published online: 27 February 2010
ꢀ Springer Science+Business Media, LLC 2011
Abstract A series of new coumarin derivatives 4 con-
taining a chalcone moiety were synthesized by condensation
of 3-acetyl-4-hydroxycoumarin 1 with aryl or heteroaryl
aldehydes 2 in the presence of piperidine in chloroform. The
interaction of 3-formyl-4-chloro-coumarin 3 with nitrogen
compound nucleophiles are described and lead to the cor-
responding substituted chromen[4,3-c] pyrazol-4-ones 5.
The structures of the obtained compounds were established
on the basis of IR|1D|2D NMR, while crystal structure of
3-acetyl-4-hydroxy coumarin 1 was determined using X-ray
diffraction and further were evaluated for possible antibac-
terial and antioxidant activities. The coumarinic chalcone 4d
has been found to be the most active (IC50 = 2.07 lM) in
this study.
Abbreviations
LTR
TPA
Long terminal repeat
Two photon absorption
AIDS Acquired immunodeficiency syndrome
Introduction
Coumarin derivatives constitute an important class of
compounds with a wide range of biological properties
(Brown et al., 1982; O’Kennedy and Thornes 1997;
Fylaktakidou et al., 2004) and in particular, they are
important as photochemotherapeutic agents that are used
to treat a variety of skin diseases (Dall’Acqua et al.,
1995). They have been also found to exhibit antitumor,
antioxidant (Bordin et al., 1995) and antiinflammatory
(Mahidol et al., 2004) activities. Some phenyl coumarins
and chalcones have been proposed as suppressors of LTR-
dependent transcription, but the mechanism of action
has not been fully characterized (Borrow et al., 2004)
(?)-Calanolide A, a natural dipyranocoumarin is also
currently undergoing anti-AIDS clinical trials (Hatano
et al., 2003). Recent research suggests that the connection
of a chalcone moiety with the coumarin ring appears quite
promising for the synthesis of derivatives with enhanced
TPA cross-sections (Li et al., 2007). In the course of our
continuing interest on the synthesis of coumarin deriva-
tives including 4-hydroxyl coumarin with antibacterial
and antioxidant activities (Carmen et al., 2008; Antonio
et al., 2008; Antonio et al., 2006); we have extended our
research to the synthesis and biological evaluation of new
coumarinyl chalcones and substituted chromen[4,3-c]
pyrazol-4-ones as antibacterial and antioxidant agents.
This synthesis using 3-acetyl-4-hydroxycoumarin 1 and
Keywords Coumarin derivatives ꢀ Crystal structure ꢀ
Chalcone
N. Hamdi (&)
Borj Cedria Higher Institute of Sciences and Technology,
Environment Touristic Road of Soliman B.P. 95.,
2050 Hammam-Lif, Tunisia
e-mail: naceur.hamdi@isste.rnu.tn; hamdi_naceur@yahoo.fr
C. Fischmeister
CNRS-UMR ‘‘Sciences Chimiques de Rennes’’, Universite
´
´
Rennes 1, Laboratoire Catalyse et Organometalliques,
263 avenue du general Leclerc Batiment 10C,
´ ´
ˆ
35042 Rennes cedex, France
M. C. Puerta ꢀ P. Valerga
´
Departamento de Ciencia de los Materiales Ingenierıa
Metalurgica y Quımica Inorganica, Facultad de Ciencias,
´
´
´
´
Universidad de Cadiz, Campus del Rıo San Pedro,
11510 Puerto Real, Spain
´
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Med Chem Res (2011) 20:522–530
523
OH
O
Cl
O
O
CH3
H
O
O
O
1
3
Scheme 1 3-Acetyl-4-hydroxycoumarin and 4-chloro-3-formyl cou-
marin target molecules
4-chloro-3-formyl coumarin 3 systems via an easy and
rapid method, which improves the yields to a significant
level, is reported here (Scheme 1).
Results and discussion
For acetylation of the 4-hydroxycoumarin to give 3-acetyl-
4-hydroxycoumarin 1, the method of (Dholakia et al.,
1968) was employed using glacial acetic acid as acetylating
agent in the presence of POCl3.
Fig. 1 Unit cell view of the crystal down c-axis showing intermo-
lecular and intramolecular hydrogen bonds (dashed lines)
The reaction was rapid, without involving any kind of
competition from the intramolecular condensation of
4-hydroxycoumarin (Hamdi et al., 1992).
Table 1 Hydrogen-bonding geometry
˚
˚
Donor_Acceptor (A)
Intermolecular hydrogen bond
H_Acceptor (A)
1
This compound was characterized by IR, H and 13C
NMR. The 1H NMR shows that aromatic proton arrow as a
multiplet between 7.24 and 8.01 ppm. A singlet at 2.74 ppm
was assigned to methylic proton whilst the OH signal
appeared at 17.72 ppm. This very high value of the chem-
ical shift might be explained by only an intermolecular
hydrogen bond (Palinko et al., 1995; Hamdi et al., 1993;
David and Michael, 2001). Consequently, we became con-
cerned about the identification of this compound and
therefore carried out an X-ray structure determination, in
order to establish unambiguously the structure.
O3A_O2A
O3B_O2B
2.445(4) H8_O2A
2.436(4) H8B_O2B
2.920(4) H8_O3A
1.412(3)
1.502(50)
2.735(3)
O3A_O3A [-x - 1,
-y ? 1, -z]
O3B_O3B [-x, -y ? 1, 2.923(4) H8B_O3B
-z ? 1]
2.576(43)
The compound crystallizes in the triclinic space group
P-1 with two molecules in the unit cell. The 3-acetyl-4-
hydroxycoumarin molecules are arranged separately in two
quasi-perpendicular layer planes. The layer planes are
˚
stacked parallel to each other at 3.354 A apart and an
alternating (ABAB-) stacking sequence is observed as in
graphite structure (Fig. 1). In each layer the molecules are
separated and this may make the material desirable as a
solid lubricant.
Fig. 2 ORTEP diagram of the compound 1 at 50% probability
The structure of compound 1 exhibits also intramolec-
ular hydrogen bond of the type O–HꢀꢀꢀO (Table 1; Fig. 2).
The intramolecular hydrogen bond between the hydroxyl
group and the ketonic oxygen is O3A–H8AꢀꢀꢀO2A in
‘‘molecule 1 (O3B–H8BꢀO2B in molecule 2)’’ which has a
hydroxycoumarin 1 with various aryl or heteroaryl alde-
hydes in the presence of piperidine in chloroform.
In conventional methods for chalcone synthesis, the
time for completion of reactions is very long, ranging from
24 to 36 h at rt (Joshi et al. 2003) A small alteration in
reaction conditions, using chloroform as solvent with a
mild organic base, for example, piperidine, reduced the
˚
˚
length of 1.412 A (1.502 A).
The synthesis of coumarinic chalcones was achieved in
one step using a new pathway by refluxing 3-acetyl-4-
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Med Chem Res (2011) 20:522–530
OH
O
OH
OH
O
O
O
Scheme 2 Synthesis of
coumarinic chalcones 4
H
a
b
+
R
Ar
O
O
O
O
O
1
2
4
Reagents and conditions: (a) glacial acetic acid, POCl3 (b) CHCl3, piperidine, 80ºC.
reaction time in most cases, to 1–1.5 h. Moreover, the
isolation of product 4 was facilitated.
The coumarinic chalcone structure was deduced from an
analysis of HMBC spectra which indicate that both ethylenic
protons correlate with ketone carbon atom via ‘‘J2 and J3’’
coupling constant, and C3, respectively. Like 3-acetyl-4-
hydroxycoumarin compounds 4a–i can exist in several tau-
tomeric forms. The most stable tautomers, A and B, are
stabilized by intramolecular hydrogen bonding (Scheme 3).
In contrast to 3-acetyl-4-hydroxycoumarin the cou-
marinic chalcone A is more stable for all compounds 4.
The electronic absorption spectra of compounds 4a–i
displayed absorption maxima in the visible region from
400 to 800 nm depending on the nature of acceptor group.
In the case of the coumarinic chalcones, 4a–e and 4f–i, the
introduction of groups as an electron acceptor resulted in a
large bathochromic shift relative to the coumarinic chal-
cones 4c and 4d having methoxylic and methyl groups.
Especially coumarinic chalcone 4h containing naphtyl
group showed remarkable shift in lambda max towards
longer wavelength (Table 2).
All the products were obtained as solids and their purity
was checked by thin layer chromatography (eluant: hexane/
ethyl acetate, 1/1. v/v). The reaction studied and the
products obtained are depicted in Scheme 2.
All the synthesized compounds have been characterized
on the basis of their physical data and spectral analysis.
4a–i
Ar
a
b
c
d
e
f
C6H5
pF–C6H4
pOCH3C6H4
pMe C6H4
pBr C6H4
pNO2 C6H4
3,4,5 OCH3 C6H2
C10H7
g
h
i
pN(Me)2 C6H4
All the coumarinic chalcones considered absorb in the
same spectral region, but the shape of their absorption
spectra differs with the nature of the tether. The magnitude
of the molar absorption coefficient is typical of p–p tran-
sitions. The experimental UV–visible spectra of coumari-
nic chalcones 4a–i diluted in chloroform were obtained and
are shown in Fig. 3.
The IR spectra of compounds, 4a–i, showed bands
resulting from the OH, (ketone) C=O and (lactone) (C=O)
stretching in the region 3305–3320 cm-1, 1670–1710 cm-1
and 1720–1750 cm-1, respectively, in all the cases. All these
evidences were supportive of the formation of compounds
4a–i.
The 1H NMR spectra of 4c in DMSO-d6 showed singlet
at d 3.84 for three protons attributed to the methoxylic
group, signal at d 19.05 ppm for the OH proton whilst the
rest of the 10 protons appeared as multiplets between d
6.91 and 8.33.
The 13C NMR spectrum of 4c in ‘‘DMSO-d6’’ showed
two downfield signals at d 162.42(ketone ‘‘C=O’’) and d
157.41(lactone ‘‘C=O’’) as well as one up field signal at
d55.48 (OCH3). Measurement of the spectrum using the
DEPT technique showed that the HC carbons of the cou-
marinic ring appear at d 134.5, 131.3, 125.6 and 124.11.
A range of activities have been claimed to arise from the
fusion of some coumarins with pyrazole ring (Malhotra
et al., 1988; Evans et al., 1996). Moreover, recently
substituted pyrazoles have been used as analytical reagents
in the complexation of transition metal ions (Artamonova
et al., 1983; Kalaj et al., 1987).We have attempted to
exploit this behaviour of aldehyde groups in 4-chloro-3-
formyl coumarin with the purpose of finding a satisfactory
route by which to synthesize substituted chromen[4,3-c]
pyrazol-4(1H)-ones bearing hydrogen or alkyl groups on
N2. The compound 3 was prepared from 4-hydroxycoum-
arin under Vilsmeier conditions (POCl3/DMF). Literature
(Eggenweiler et al., 2001; Tabakovic’ and Tabakovic’,
H
H
Scheme 3 Tautomeric forms
of compounds 4
O
O
O
O
O
O
R
R
O
O
A
B
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Med Chem Res (2011) 20:522–530
525
hydrazines were allowed to react in ethanol at reflux, for
2 h and led to the quantitative formation of the substituted
chromen[4,3-c] pyrazol-4-ones, 5, in short times and in
high yields (67–89%). The reaction pathway is depicted in
Scheme 4.
Table 2 Spectral properties of the coumarinic chalcones 4a–i in
chloroform
Compounds 4a–i
kmax (nm)
e mol-1 dm3 cm-1
a
b
c
d
e
f
495
471
450
474
465
485
463
475
480
30500
23000
16100
8330
The proposed structures of the new substituted chro-
men[4,3-c] pyrazol-4(1H)-ones, 5, were validated by the
spectral data, which were consistent with available litera-
ture data for similar substitution patterns in coumarin
derivatives (Kadaba et al., 1984; Ivanov et al., 1992).
14711
23100
14500
28500
7200
1
g
h
i
The H NMR spectra of the compounds 5a showed the
expected signals: an exchangeable NH proton and the
typical doublet for coumarin 9-H in the range d
7.96–8.20 ppm, validated by the 13C spectral pattern of the
coumarin ring. All the chromen[4,3-c] pyrazol-4(1H)-ones
5a–c displayed IR stretching at 1711–1742 cm-1 associ-
ated with lactone carbonyl groups, also confirmed by the
signals appearing between d 155 and 162 ppm in the 13C
NMR spectra.
The derivatives 5a can exist in tautomeric forms a, b
and c, depicted in Fig. 4.
Annular tautomerism is common in heteroaromatic
compounds and has been widely studied, but literature
relating to the chromen[4,3-c] pyrazol-4(1H)-ones nucleus
gave poor attention to this tautomerism problem (Kalaj
et al., 1987; Eggenweiler et al., 2001). The authors of the
cited references assign two opposite tautomeric structures to
the same compound and the 1H NMR spectroscopic data for
this product are not very detailed. In our case, the b tautomer
can be ruled out because all the IR spectra of the chro-
men[4,3-c] pyrazol-4(1H)-ones 5a–c, recorded in chloro-
Fig. 3 Absorption spectra of the coumarin chalcones in chloroform
solution
form solution, showed lactone bands at 1704–1712 cm-1
,
1981; Heber et al., 1995; Oh and Park, 1994) claims the
formation of mixture of carbaldehyde 3 and 4-chloro-
coumarin without any indication of their ratio.Hantsch-
mann et al. (1992) reported that the reaction mixture
contained a mixture of 4-chlor-3-coumarincarbaldehyde 3
(65%) with 4-chlorocoumarin as a side product (20%).
After further optimization, we could obtain up to 95% of
the carbaldehyde 3. TLC was used to monitor the progress
of the reaction. The spectroscopic data MS, IR and
1H-NMR are in agreement with the structure of 4-chloro-
3-formylcoumarin. The heating of 3-formyl-4-chloro-
coumarin 3 and the appropriate nitrogen compounds
confirmed by typical carbonyl patterns in the 13C NMR
spectra. 1H and 13C NMR analyses show that compounds 5
exist as the tautomeric containing a proton on the N atom.
The reported derivatives, 4a–i, were tested for their
antioxidant and antibacterial activities.
DPPH radical scavenging activity
There is an increasing interest in antioxidants, particularly
in those intended to prevent the presumed deleterious
effects of free radicals in the human body, and to prevent the
R
Scheme 4 Synthesis of
chromen[4,3-c] pyrazol-4(1H)-
ones 5
N
OH
Cl
O
N
H
POCl3
DMF
RNHNH2
Ethanol, Et3N
O
O
O
O
O
O
3
5a-c
R
H
CH3
Ph
5
a
b
c
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Med Chem Res (2011) 20:522–530
N
N
N
N
Fig. 4 Tautomeric forms of
chromen[4,3-c] pyrazol-4(1H)-
ones 5
HN
N
O
O
O
OH
O
O
α
β
γ
deterioration of fats and other constituents of food stuffs. In
both cases, there is a preference for antioxidants from nat-
ural rather than from synthetic sources. There is therefore a
parallel increase in the use of methods for estimating the
efficiency of such substances as antioxidants.
noted that in all cases, any residual (yellow) colour from
the reduced form or any non specific absorbance from the
sample has to be taken into account in defining the
‘‘endpoint’’ of the titration, or the 50% point, this IC50
parameter also has the drawback that the higher the
antioxidant activity, the lower is the value of IC50. This is a
disadvantage particularly when results are presented
graphically as a bar chart even if the same data are
available in numerical forms (Fig. 5).
One such method that is currently popular is based upon
the use of the stable free radical diphenylpicrylhydrazyl
(DPPH). The purpose of our study is to examine the use of
the parameter ‘‘IC50’’ (equivalent concentration to give
50% effect) which is currently used in the interpretation of
experimental data from the method. When a solution of
DPPH is mixed with that of a substance that can donate a
hydrogen atom, then this gives rise to the reduced form
with the loss of this violet colour (although there would be
expected to be a residual pale yellow colour from the
picrylgroup still present). Representing the DPPH radical
by Z• and the donor molecule by AH, the primary reaction
is
The IC50 values exhibited by coumarinic chalcones 4 are
summarized in the following table:
Compounds 4a–i
EC50 (mM) 10-3
4a
4b
4c
4d
4e
4f
2.15
2.23
2.81
2.07
2.25
3.21
2.29
2.39
2.39
Zꢁ þ AH ¼ ZH þ Aꢁ
ð1Þ
where ZH is reduced form and A• is free radical produced
in this first step. This latter radical will then undergo fur-
ther reactions which control the overall stoichiometry, that
is, the number of molecules of DPPH reduced (decolou-
rized) by one molecule of the reductant.
4g
4h
4i
Compounds 4a and 4d presented potent antioxidant
activity components. While coumarinic chalcone 4f was
inactive even at a concentration of 3.21 Lm
The reaction [1] is therefore intended to provide the link
with the reactions taking place in an oxidizing system, such
as the autoxidation of a lipid or other unsaturated sub-
stance; the DPPH molecule Z• is thus intended to represent
the free radicals formed in the system whose activity is to
be suppressed by the substance AH•. Representing the
DPPH radical by Z• and the coumarinic chalcones by ROH,
the initial reaction is then Z• ? ROH = ZH ? RO• [1],
the free radical RO• evidently then reacts with another
molecule of the same kind that was produced by a parallel
reaction to (1)
ROꢁ þ ROꢁ ¼ RO ꢂ OR
ð2Þ
This therefore leads to the observed reduction of two
molecules of DPPH by two molecules of coumarinic
chalcones, that is, a 1:1 stoichiometry. One parameter that
has been introduced recently for the interpretation of the
results from the DPPH method is the efficient
concentration or IC50 value (otherwise called the IC50
value), this is defined as the concentration of substrate that
causes 50% loss of the DPPH activity (colour), and it is
corresponding to the endpoint of the titration. It should be
Fig. 5 DPPH radical scavenging activities of coumarinic chalcones
4a–i
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Med Chem Res (2011) 20:522–530
527
The reaction conditions are simple, since they are not
sensitive to oxygen and water, which make it easy to
operate at reflux. We have shown a new synthetic method
to synthesize substituted chromen[4,3-c] pyrazol-4-ones 5
in good yields. The flexibility of this methodology should
provide access to many various coumarinyl pyrazoles. All
these compounds are observed to exhibit excellent absor-
bance properties. The DPPH method has been widely
applied for estimating antioxidant activity in recent years,
but its applications should be carried out bearing in mind
the basis of this method, and the need wherever possible to
establish the stoichiometry for the quenching reaction, so
that the antioxidant activity may be related to the structure
of the substrate molecule.
Table 3 Antibacterial screening for compounds 4a–i and 5a–c
Compounds Concentration (mg/
disc)
Inhibition zone, diameter
(mm)
4a
4b
4c
4d
4e
4f
1
1
1
1
1
1
1
1
1
1
1
1
1
10
13
14
13
9
16
11
8
4g
4h
4i
11
20
14
17
19
15–20
4j
5a
5b
5c
Our chemical study on the synthesis of coumarinic
derivatives allowed us to obtain and identify 12 com-
pounds. The presence of coumarin nucleus gives to this
species an important pharmacological and therapeutic
interest. Antioxidant activities of these compounds against
the stable free radical DPPH show that these species are
good source of compounds that could help to increase the
overall antioxidant capacity of an organism.
Gentamycin 1
Compound 4d proved to be the most active compound in
this study.
Comparing the activity of compounds 4a–c, 4e and 4g–i
it was concluded that there was no substantial difference
when we changed the nature of the group R.
Experimental section
Antibacterial activities
General procedures
New coumarinic chalcones derivatives 4 were also screened
in vitro for their antimicrobial activities against Gram-
positive bacteria, Staphylococcus aureus (NCTC-7447),
using the paper disc diffusion method for the antibiotic
sensitivity technique (NCCLS 1997). The tested compounds
were dissolved in CHCl3 to obtain 1 mg/ml solution. The
inhibition zones of microbial growth produced by different
compounds were measured in millimetres after 24 h of
incubation at 37ꢁC. The results are summarized in Table 3.
The antibacterial activities for compounds 4a–i and 5a–
c show that the activity against bacteria is moderate, but in
addition, it was clearly demonstrated that this kind of
compounds could be an antibacterial agent, it’s activity
depends on its chemical composition. The antibacterial
activities for these compounds were evaluated. The mod-
erate active antibacterial effects observed showed that this
kind of compounds could be an antibacterial agent.
Flash chromatography was carried out on 0.04-0.063 mm
(Merck) silica gel, thin layer chromatography was carried out
on aluminium backed silica plates by Merck and plates were
revealed using a UV 254 light. 1H NMR (300 MHz) and 13
C
NMR (75 MHz) spectra were recorded on a Varian VXR 300
instrument at 293ꢁK in CDCl3 or DMSO d-6. Spectra were
internally referenced to TMS. Peaks are reported in ppm
downfield of TMS. Multiplicities are reported as singlet (s),
doublet (d), triplet (t), quartet (q), some combinations of
these were made by DEPT editing of the spectra. The IR-
spectra were recorded on a Philips Analytical PU 9800
spectrometer. The melting points of compounds were
determined in open glass capillaries in a paraffin bath and are
uncorrected. The MS-spectra were recorded on an AEI MS
902 S electron ionization spectrometer (EI = 70 eV). The
absorption spectra were measured on a Beckmann K25
spectrophotometer. Elemental microanalysis was obtained
by the microanalytical laboratory services, university of
Almeria in Spain.
Conclusion
In summary, we developed a new and simple method for
the easy synthesis of new coumarinyl chalcones derivatives
through a condensation reaction, which are easier.
Our improved synthesis of coumarinyl chalcones was
effective in terms of time and yields.
Materials
1,1-Diphenyl-2-picrylhydrazyl (DPPH) was obtained from
Sigma. All other chemicals were of analytical grade purity.
The 4-hydroxycoumarin, aromatic aldehydes and nitrogen
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Med Chem Res (2011) 20:522–530
Characterization of newly synthesized coumarinic
binucleophilic were purchased from Fluka, Staphylococcus
aureus (NCTC-7447) (National Collection of Type Cul-
tures 7447)
chalcones
3-((2E)-3-(phenyl)prop-2-enoyl))-4-hydroxy-2(H)-
chromen-2-one (4a)
3-Acetyl-4-hydroxycoumarin (1)
Solid (yield 79%), mp = 145ꢁC, IR: m 3168 (–OH), 1622
([C=O), 1577 (C=C), 1028(s) (sym) (C–O–C); 1H NMR: d
To a solution of 4-hydroxy-2H-chromen-2-one (3 g,
1.86 mmol) in acetic acid (16 ml) was added phosphorus
oxychloride (5.6 ml). The mixture was heated at reflux for
30 min. After cooling, the precipitate was collected and
recrystallized from ethanol to give 3-acetyl-4-hydroxy-2H-
chromen-2-one as white needles. Yield 2.7 g (90%); mp
135ꢁC. IR spectrum, m cm-1: 3185 (OH); 1705 (CO); 1700
´
(ppm) 7.2–8.5 (m, 10H, Ar–H ? Hethy), 18.56 (s,1H,
OH). 13C NMR (ppm): 181.5 (CO); 166 (C4); 163.5 (C2);
116.3–136.2 (Carom); 154.8 (=-Ph); 146.0 (=-CO).
C18H1204 calc. C 73.97 H 4.10 O 21.92; found C 73.90 H
4.10 O 21.80.
1
(O–CO lactone). H NMR spectrum (CDCl3). d ppm: 2.72
(3H, s, CH3); 7.98 (1H, s, H-5); 7.95 (1H, dd, 3J7.8 = 8.35,
4J6.8 = 1.2, H-8); 7.1–7.4 (2H, m, H-6 ? H-7); 17.69 (1H,
s, OH). 13C NMR spectrum (CDCl3), d ppm: 29.9 (CH3);
178.5 (CO); 159.8 (C-4); 154.6 (C-2); 101.26 (C-3);
115.0–136.0 (Carom). Mass spectrum, m/z (I, %): 204 [M]?
(100); 189 (74); 161 (43).
3-((2E)-3-(4-fluorophenyl)prop-2-enoyl)-4-hydroxy-2(H)-
chromen-2-one (4b)
Solid (yield 80%), mp = 178ꢁC, IR: m 3268 (–OH), 1625
([C=O), 1575 (C=C), 1025(s) (sym) (C–O–C); 1H NMR: d
´
(ppm) 6.86–7.73 (m, 10H, Ar–H ? Hethyl), 19.58 (s,1H,
OH). 13C NMR (ppm): 192.2 (CO); 173.9 (C4); 172.2 (C2);
´
103.2–167.8 (Carom ? Cethyl). C18H1104 F calc. C 69.67 H
4-Chloro-3-coumarincarbaldehyde (3)
3.54 O 20.64 found C 69.70 H 3.60 O 20.70.
To a stirred mixture of 4-hydroxycoumarin (9.72 g,
0.06 mol) in anhydrous DMF (46.2 ml, 0.6 mol) were added
dropwise POCl3 (27.6 g, 0.18 mol) at -10ꢁ to -5ꢁC. The
reaction mixture was then stirred for 1 h at room temperature
and heated and stirred for 2 h at 60ꢁC. After the reaction was
completed, the mixture was poured onto crushed ice (200 g)
under vigorous stirring. After storing the mixture overnight
at 0ꢁC the pale yellow solid was collected by filtration and
washed successively with aqueous Na2CO3 (5%) and water,
and then was air-dried. 4-Chlorocoumarin (2.50 g, 20%) was
separated by Soxhlet extraction as the second product.
Recrystallization from acetone gave 8.99 g (72%) of 2 as a
pale yellow powder with mp 115–120ꢁC; 1H NMR d 10.39
(1H, s, CH=O),8.19–7.40 (4H, m, ar–H); 13C NMR d 186.81
(11-C), 158.44 (2-C), 153.28 (4-C), 153.27 (9-C),135.68
(7-C), 127.65 (5-C), 125.56 (6-C), 118.39 (10-C), 118.22
(3-C), 117.20 (8-C); IR m 2920, 2874, 1720, 1702, 1603,
1587, 1541 cm-1; MS m/z: 208 (MH?, 11), 182 (31), 180
(100), 154(31), 152 (91), 124 (20), 101 (11).
3-((2E)-3-(4-methoxyphenyl) prop-2-enoyl)-4-hydroxy-
2(H)-chromen-2-one (4c)
Solid (yield 70%), mp = 135ꢁC, IR: m 3250 (–OH), 1620
([C=O), 1574 (C=C), 1025(s) (sym) (C–O–C); 1H NMR: d
(ppm) 3.84 (s, 3H, OCH3), 6.91–8.33 (m, 10H, Ar–
H ? Hethy), 19.05 (s,1H, OH); 13C NMR (ppm): 55.4
´
(OCH3); 162.4(CO); 147.4 (C=-Ph); 135.6 (=-CO);
114.4–134.5 (Carom). C19H1405 calc. C 70.80 H 4.34 O
24.84 found C 70.70 H 4.40 O 24.80.
3-((2E)-3-(p-tolyl) prop-2-enoyl)-4-hydroxy -2(H)-
chromen-2-one (4d)
Solid (yield 75%), mp = 138ꢁC, IR: m 3368 (–OH), 1627
([C=O), 1575 (C=C), 1030(s) (sym) (C–O–C); 1H NMR: d
´
(ppm) 2.3 (s, 3H CH3), 7.2–8.2 (m, 10H, Ar–H ? Hethy),
17.82 (s, 1H, OH); 13C NMR (ppm): 30.0 (OCH3);
178.4(CO); 154.6 (C4); 135.6 (C2); 101.3–135.9
(Carom).C19H1404 calc. C 74.50 H 4.57 O 20.91, found C
74.40 H 4.60 O 20.90.
General procedure for the preparation of 3-((2E)-3-(4-
substituted phenyl prop-2-enoyl)-4-hydroxy-(2H)-
chromen-2-ones (4a–i)
3-[(2E)-3-(4-bromophenyl)prop-2-enoyl]-4-hydroxy-2(H)-
chromen-2-one (4e)
3-Acetyl-4-hydroxyxcoumarin (0.031 mol) and the substi-
tuted aromatic aldehyde (0.03 mol) were dissolved in
30 ml of chloroform. A catalytic amount of piperidine
(0.02 mol) was added and the reaction mixture was
refluxed for 1.5 h. The chloroform was distilled out and the
residue was washed with methanol.
Solid(yield 85%), mp = 182ꢁC, IR: m 3250 (–OH), 1625
([C=O), 1578 (C=C), 1030(s) (sym) (C–O–C); 1H NMR: d
´
(ppm) 6.26–7.84 (m, 10H, Ar–H ? Hethy), 19.58 (s, 1H,
OH); 13C NMR (ppm): 191.2 (CO); 172.8 (C4); 172.1
123

Med Chem Res (2011) 20:522–530
529
´
(C2); 102.2–168.8 (Carom ? Cethyl). C18H1104Br calc. C
After addition of the reagent was complete a yellowish
precipitate formed rapidly. The precipitate was filtered off
and recrystallized from ethanol and then twice more from
DMF to give fine colourless needles.
58.22 H 2.96 O 17.25, found C 58.30 H 2.90 O 17.30.
3-((2E)-3-(4-nitro-phenyl) prop-2-enoyl)-4-hydroxy-2(H)-
chromen-2-one (4f)
3-Chromeno[4,3-c]pyrazol-4(1H)-one 5a
Solid (yield 82%), mp = 147ꢁC, IR: m 3251 (–OH), 1715
1
([C=O), 1570 (C=C), 1075 (sym) (C–O–C); H NMR: d
´
(ppm) 6.80–8.14 (m, 10H, Ar–H ? Hethyl), 19.50 (s, 1H,
Solid (Yield 67%), mp = 183ꢁC; IR: m 3368 (–NH),
1716(s) (lactone [C=O), 1577 (C=C), 1018(s) (sym) (C–
1
OH); C18H1106 N calc. C 64.1 H 3.26 N 4.15 O 24.48,
found C 64.10 H 3.30 N 4.10 O 24.50.
´
O–C); H NMR: d (ppm) 7.40 (s, 1H, Hethy), 7.96–8.20
(m, 4H, Ar–H), 13.2 (s, 1H, NH); 13C NMR (ppm): 158.2
(C2); 102.1 (C3); 149.2 (C4); 146.7 (C=N); 122.1–151.2
(Carom). C10H502N2 calc. C 64.52 H 3.22 N 15.05 O 17.21,
found C 64.60 H 3.3 N 15.2 O 17.1.
3-((2E)-3-(3,4,5-trimethoxy-phenyl)prop-2-enoyl)-4-
hydroxy-2(H)-chromen-2-one (4g)
Solid (yield 83%), mp = 193ꢁC, IR: m 3368 (–OH),
1716(s) ([C=O), 1577 (C=C), 1018(s) (sym) (C–O–C); 1H
1-Methylchromeno[4,3-c]pyrazol-4(1H)-one 4b
´
NMR: d (ppm) 6.90–7.96 (m, 8H, Ar–H ? Hethyl), 19.50
Solid (Yield 75%), mp= = 184ꢁC; IR: m 1720(s) (lactone
[C=O), 1578 (C=C), 1020(s) (sym) (C–O–C); 1H NMR: d
(s, 1H, OH); 13C NMR (ppm): 191.6(CO); 59.7 (O-CH3);
59.8 (O–CH3). 60.0 (O–CH3) 103.47–153.3 (Carom); 164.8
(C4); 163.2 (C2); 102.8 (C3). C21H1807 calc. C 65.96 H
4.71 O 29.31, found C 65.90 H 4.80 O 29.30.
´
(ppm). 3.5 (s, 3H, CH3), 6.4 (s, 1H, Hethy); 7.13–7.45 (m,
Ar–H), 13C NMR (ppm): 157.2 (C2); 112.2 (C3); 144.6
´
(C4); 140.6 (Cethy); 122.1–151.2 (Carom). C11H702 N2 calc.
C 66 H 4 N14 O 16 found C 66.1 H 4.2 N14.1 O 16.2.
3-[(2E)-3-(naphthyl) prop-2-enoyl]-4-hydroxy-2(H)-
chromen-2-one (4h)
1-Phenyl chromeno[4,3-c]pyrazol-4(1H)-one (5c)
Solid (yield 80%), mp = 185ꢁC, IR: m 3365 (–OH),
1718(s) ([C=O), 1578 (C=C), 1019(s) (sym) (C–O–C); 1H
Solid (Yield 89%), mp = 185ꢁC; IR: m 1718(s) (lactone
[C=O), 1579 (C=C), 1022(s) (sym) (C–O–C); 1H NMR: d
´
NMR: d (ppm) 6.75–7.92 (m, 13H, Ar–H ? Hethy), 19.40
(ppm): 7.13–7.45 (m, Ar–H), 7.2 (s, 1H, Hethy) 13C NMR
´
(s, 1H, OH); 13C NMR (ppm): 192.6 (CO); 102.47–154.2
(Carom); 164.6 (C4); 164.2 (C2); 101.8 (C3).C22H1404 calc.
C 77.20 H 4.10 O 18.71, found C 77.10 H 4.10 O 18.70.
´
(ppm): 158.2 (C2); 112.4 (C3); 144.2 (C4); 138.6 (Cethy);
122.1–151.2 (Carom). C16H1002 N2 calc. C 73.28 H 3.81 N
10.68 O 12.22, found C 73.2 H 3.7 N 10.6 O 12.1.
3-((2E)-3-(4-nimethylamino-phenyl) prop-2-enoyl)-4-
hydroxy-2(H)-chromen-2-one (4i)
The experimental procedure for the antioxidant
evaluation
Solid (yield 85%), mp = 192ꢁC, IR: m 3125 (–OH),
1712(s) ([C=O), 1550(C=C), 1028 (sym) (C–O–C); 1H
NMR: d (ppm) 2.9 (s, 6H, CH3), 6.54–8.1 (m, 10H, Ar–
The DPPH radical scavenging capacity of the obtained
coumarinic derivatives was measured from the bleaching
of purple coloured ethanol solution of DPPH. Method
described by Hatano et al. (1988) was used. Half a milli-
litre of each sample concentration was mixed with an equal
volume of DPPH ethanolic solution. After incubation for
30 min in darkness at 25ꢁC, absorbance was read at
520 nm wavelength. A mixture of 0.5 ml of DPPH solution
and 0.5 ml of ethanol was taken as a blank (absorbance is
equal to zero). Inhibitory concentration (IC50) values
denoting the concentration (microgram of natural sub-
stance per millilitre of ethanol) required to scavenge 50%
of DPPH radicals were calculated. All measurements were
performed in triplicate. Results were expressed in inhibi-
tion percentage versus sample concentrations (mg/ml) at
30 min.
H ? Hethy), 18.62 (s, 1H, OH); 13C NMR (ppm): 191.0
´
(CO); 40.1 (CH3–N); 102.8–142.9 (Carom); 179.2 (C4);
162.2 (C2); 102.8 (C3).C20H1704 N calc.C 71.64 H 5.1 N
4.17 O 19.10, found C 71.60 H 5.10 N 4.20 O 19.30.
Synthesis of aryl (alkyl) chromeno[4,3-c]pyrazol-
4(1H)-one 5
The aldehyde 3 (0.41 g, 2 mmol) dissolved on boiling in
ethanol (10 ml) and was then cooled to 15–20ꢁC. A solu-
tion prepared from aryl or alkylhydrazine hydrochloride
(0.29 g, 2 mmol) and triethylamine (4 mmol) in 60%)
ethanol (10 ml) was slowly added dropwise so that the
temperature of the reaction mixture did not exceed 25ꢁC.
123

530
Med Chem Res (2011) 20:522–530
Eggenweiler M, Gassen M, Rochus J, Poeschke O, Wolf M (2001)
Merck Patent Gmbh, Germany, PCT Int. Appl. (see: Chem Abstr
134:331–619)
The experimental procedure for the antibacterial
evaluation
Evans JM, Geen GR, Vong AK (1996) In: McKillop A (ed) Pyrans
and their benzo derivatives: applications. comprehensive het-
erocyclic chemistry II. Pergamon, Oxford, pp 469–484
Fylaktakidou KC, Hadjipavlou-Litina DJ, Litinas KE, Nicolaides DN
(2004) Natural and synthetic coumarin derivatives with anti-
inflammatory/antioxidant activities. Curr Pharm 10:3813–3833
Hamdi N, Sakellariou R, Speziale V (1992) Antibacterial, antifungal
and cytotoxic properties of novel N-substituted sulfonamides
from 4-hydroxycoumarin. J Heterocycl Chem 29:1817
Hamdi M, Reine S, Patrick G, Vincent (1993) Reaction of amines on
3-ureidomethylenecoumarins. A new route to N-(methylene-4-
oxocoumarinyl) amines. J Hetercycl Chem 30:1155–1157
Hantschmann A, Liebigs M, Steinfuhrer T, Weissenfels M (1992)
Ann Chem 23
The obtained products are dissolved in a suitable solvent,
the determined quantities of this solution are put on the disc
(example: 1 mg/disc, it is enough to dissolve 1 mg of the
sample in 0.1 ml of CHCl3), then the solvent is evaporated
and the product remained. This prepared disc is called a
treated disc. The second disc must be prepared: it is a disc
of control (the solvent used can obtain active products, to
ensure, the same volume has to be taken to dissolve the
sample. The third disc must contain gentamycin, and the
latter is for comparison. Then we put all three discs on
the surface of gel containing bacteria, and then we close it
at 37ꢁC for 24 h, and we measure the diameters of the clear
zones.
Hatano T, Kagawa H, Okuda T, Yasuhara T (1988) Antioxidant
activity and volatile components of Egyptian Artemisia judaica
L. Chem Phar Bull 36:2090–2097
Hatano T, Ito H, Tanuma S, Uchiumi F, Yoshida T (2003)
Transcriptional suppression of the HIV promoter by natural
compounds. Antiviral Res 58:89–98
Heber D, Ivanov IC, Karagiasov SK (1995) J Heterocycl Chem 32:
505
Acknowledgements This work was carried out with financial aid of
both Tunisian Ministry of Higher Education and Scientific Research
and Technology and the Spanish Agency of International Cooperation
through projects (A/9549/07and A/8302/07).
Ivanov IC, Karagiosov SK, Simeonov MF (1992) A facile synthesis
of benzopyrano[4,3-b]pyridin-5-ones. Liebigs Ann Chem
203–207
Joshi HS, Kachhadia VV, Nimavat KS, Potal KH (2003) Improved
and rapid synthesis of new coumarinyl chalcone derivatives and
their antiviral activity. J Ind Chem Soc 80:707–708
Kadaba PK, Tisler M, Stanovnik B (1984) 1,2,3-Triazole and its
derivatives: development of methods for the formation of the
triazole ring. Adv Heterocycl Chem 37:339
References
Antonio R, Hedi Z, Chaker L, Hamdi N, Mustapha S (2006) Synthesis
of some new biologically active coumarin derivates. Chem
Heterocycl Comp 3:361–366
Antonio R, Hamdi N, Mustapha S (2008) Synthesis, spectroscopic
and antibacterial investigations of new hydroxyl ethers and
Kalaj V, Kitan D, Trkovnik M (1987) Synthesis of new hetero-
cyclocoumarins from 3,4-damino-and 4-chloro-3-nitrocouma-
rins. Org Prep Proc Int 19:450–455
heterocyclic coumarin derivates.
45:1835–1842
J
Heterocycl Chem
Artamonova OS, Chernyakova IV, Malysheva LI, Pryanishnikova
NT, Savel’ev VL, Shavyrina VV, Zagorevskii VA (1983)
Synthesis and pharmacological activity of 4H-(1)-benzopyrano-
(3,4-d)-imidazol-4-ones. Khim Farm Zh 17:697–700 (see Chem
Abstr 99:158–325)
Bordin F, Chilin A, Dall’Acqua F, Guiotto A, Manzini P (1995)
Synthesis of angelicin heteroanalogues: preliminary photobio-
logical and pharmacological studies. Rod Il Farm 50:479–488
Borrow E, Barrow W, Flavin MT, Lin YM, Suling WJ, Xu ZQ,
Westbrook L (2004) Chemical resolution of ( )-calanolide A,
Li X, Zhao Y, Wang T, Shi M, Wu F (2007) Coumarin derivatives
with enhanced two-photon absorption cross-sections. Dyes
Pigments 74:108–112
Mahidol C, Ploypradith P, Sahakitpichan P, Wongbundit S, Ruchira-
wat (2004) A Highly efficient synthesis of Lamellarins K and L
by the Michael addition/ring-closure reaction of benzyldihy-
droisoquinoline derivatives with ethoxycarbonyl-b-nitrostyrenes.
Angew Chem Int 43:866–868
Malhotra S, Parmar VS, Sharma VK (1988) Synthesis of three new
dihydropyranochalcones: structural revision of crotmadine, an
antifungal constituent of Crotalaria madurensis. J Nat Prod
51:578–581
( )-cordatolide A and their 11-demethyl analogues. Bioorg Med
Chem 12:1199–1207
Brown SA, Mendez J, Murray DH (1982) The natural coumarins.
Occurrence, chemistry and biochemistry. Wiley, New York
Carmen P, Hamdi N, Pedro V (2008) Synthesis, structure, antimi-
crobial and antioxidant investigations of dicoumarol and related
compounds. Eur J Med Chem 43:2541–2548
NCCLS (1997) Methods for dilution antimicrobial susceptibility. Test
for bacteria that grow aerobically, 4th ed, Approved Standard.
NCCLS Document M7-A4. NCCLS, Villanowa, PA
O’Kennedy R, Thornes RD (1997) Coumarins: biology, applications
and mode of action. Wiley, Chichester
Dall’Acqua F, Horspool WM, Vedaldi D, Song PS (1995) Handbook
of organic photochemistry and photobiology. CRC Press, Boca
Raton, pp 1341–1350
David AL, Michael TH (2001) A C-HꢀꢀꢀO=C hydrogen bond
intramolecular hydrogen bonding in a novel semirubin. J Org
Chem 66:8402–8410
Dholakia VN, Parekh MG, Trivedi NK (1968) Improved and rapid
synthesis of new coumarinyl chalcone derivatives and their
antiviral activity. Aust J Chem 22:345–2347
Oh C, Park KP (1994) J Heterocycl Chem 31:841
Palinko I, Toroka BB, Rozsa-Tarjanyi M, Kiss JT, Tasi GY (1995)
Hydrogen bonding interactions of a-phenylcinnamic acid iso-
mers in the liquid phase studied by IR and NMR spectroscopies
and computational methods. J Mol Struct 348:57–60
Tabakovic0 K, Tabakovic0 I (1981) Chemistry of coumarins. Nucle-
ophilic substitutions of chloro-3-nitrocoumarin with hard and
soft nucleophiles. Croat Chem Acta 54:451–458
123