Chemistry of phosphorus ylides. Part 33. Synthesis and antitumor activities of some new chromenone derivatives

Article abstract of DOI:10.1002/ardp.201000341

The reaction of nucleophilic phosphacumulene and phosphallene ylides with different chromenone derivatives was investigated. Heterocycles and carbocycles of various ring sizes and heteroatom patterns such as pyrano-, oxaphosphino-, cyclopenta-, phosphoran

Full text of DOI:10.1002/ardp.201000341

442
Arch. Pharm. Chem. Life Sci. 2011, 344, 442–450
Full Paper
Chemistry of Phosphorus Ylides. Part 33. Synthesis and
Antitumor Activities of Some New Chromenone Derivatives
Soher S. Maigali, Mansoura A. Abd-El-Maksoud, and Fouad M. Soliman
Organometallic and Organometalloid Chemistry Department, National Research Centre, Dokki, Cairo, Egypt
The reaction of nucleophilic phosphacumulene and phosphallene ylides with different chromenone
derivatives was investigated. Heterocycles and carbocycles of various ring sizes and heteroatom
patterns such as pyrano-, oxaphosphino-, cyclopenta-, phosphoranylidene cyclobutane-, and
phospholo-chromenones were obtained. The antitumor (breast and liver) properties of some new
compounds were performed in vitro. Some of these compounds were more potent than the
comparative standard.
Keywords: Oxaphosphino-chromenones / Phosphacumulenes / Phosphallene / Phospholo-chromenones
Pyrano-chromenones
Received: November 8, 2010; Revised: December 23, 2010; Accepted: January 11, 2011
DOI 10.1002/ardp.201000341
Introduction
Results and discussion
Coumarins are very well known for their biological activity
[1]. They exist in a variety of forms due to the various sub-
stitutions possible in their basic structure which modulate
their biological activity [2, 3]. Coumarins possess anti-inflam-
matory, antioxidant, anti-allergic, hepatoprotective, anti-
thrombotic, antiviral, antimicrobial, and anticarcinogenic
properties. Moreover, the function of plant growth and
growth regulators depends on them, as well as the control
of respiration, photosynthesis and defense against infection
[4, 5]. Evolution developed a very long association of plant
coumarins with other organisms which many account for the
extraordinary range of their biochemical and pharmacologi-
cal activities in mammals and other biological systems.
Owing to our interest in the field of synthesis of naturally
occurring analogues containing the phosphorus moieties
[6–10], we planned to study the reactions of active phos-
phonium ylides with coumarin derivatives to prepare new
heterocycles of various ring sizes and heteroatom patterns
which present the pivotal core of many biologically or phar-
maceutically interesting compounds.
Chemistry
Now, we found that reactions 4-hydroxy-2H-chromen-2-one 1
with (N-phenyliminovinylidene)-2a or (2-oxovinylidene)-tri-
phenylphosphorane 2b afforded the corresponding 2-oxo-2H-
chromen-4-yl-1-N-phenyl-2-(triphenyl-l5-phosphanylidene) etha-
nimidoate 3a and 2-oxo-2H-chromen-4-yl(triphenyl-l5-phospha-
nylidene) acetate 3b. The reaction proceeded by addition of the
–
–
OH group of 1 to C C bond of 2. The most important features
are the disappearance of the OH group of compound 3a in its IR
and 1H-NMR spectra, which appeared in the starting material 1
at 3444 cmꢀ1 and d 12.52 ppm spectra, respectively. A signal at
13.6 ppm in its 31P-NMR spectrum was found which support a
phosphorane structure [11–13] (Scheme 1).
An efficient way for the synthesis of pyranochromones has
been developed in this work. When 3-acetyl-4-hydroxy-2H-
chromen-2-one 4 was allowed to react with the active phos-
phoranes 2a and 2b in boiling toluene for 6 h in case of 2a
and 10 h when 2b was used, the corresponding 4-methyl-2-
(phenylimino)pyrano[3,2-c]chromen-5(2H)-one 6a and 4-
methyl-2H,5H-pyrano-[3,2-c]chromene-2,5-dione 6b were
obtained, respectively, together with triphenylphosphine
oxide. Compounds 6a and 6b were formed via the formation
of the complex phosphoranes 5, followed by intramolecular
Wittig reaction affording 6a and 6b. The reaction of hexa-
phenylcarbodiphosphorane 7, with 3-acetyl-4-hydroxy-2H-
chromen-2-one 4 was studied, too. When the chromenone
4 was reacted with the phosphorane 7, 4-methyl-2,2,2-tri-
Correspondence: Fouad M. Soliman, Department of Organometallic and
Organometalloid Chemistry, National Research Centre, El-Behooth St.,
P. O. 12622, Dokki, Cairo, Egypt.
E-mail: solimanfma2@yahoo.com
Fax: þ02 333 70931
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Arch. Pharm. Chem. Life Sci. 2011, 344, 442–450
New Chromenone Derivatives
443
X
We have found that the reaction of 3-acetyl-2H-chromen-2-
one 10 with (N-phenyliminovinylidene)triphenylphosphor-
ane 2a proceeded in tetrahydrofuran at room temperature
for 10 h gave 3-methyl-1-(phenylimino)-cyclopenta[c]chro-
men-4(1H)-one 13a and 3-{1-[2,4-bis(phenylimino)-3-(tri-
phenyl-l5-phosphanylidene)cyclobutylidene]ethyl}-2H-chro-
OH
O
O
C
H
PPh3
Ph3P=C=C=X
2a, X= N-Ph
O
O
b, X= O
O
1
3a, X= N-Ph
b, X= O
–
men-2-one 14a. A [2 þ 2] cycloaddition of the C O bond of 10
–
to the ylidic C-P bond of 2a furnishes the oxaphosphetane 11
[16, 17]. The possible reaction route consists in elimination of
triphenylphosphine oxide from 11 to form the ketimine 12,
which in turn undergoes intramolecular cyclization to give
13a or cycloaddition to 2a, giving the four-membered ring
14a. The most important features in the spectroscopic data of
the chromenone 13a and phosphoranylidene-cyclobutyli-
dene 14a is that they show the m/z at 286 [M – H]þ and
31P-NMR at d 30.03 ppm, respectively. When 3-acetylchrome-
none 10 was allowed to react with (2-oxovinylidene)triphe-
nylphosphorane 2b in boiling toluene for 8 h, the
corresponding compounds, 3-methylcyclopenta[c]chro-
mene-1,4-dione 13b and 2-[1-(2-oxo-2H-chromen-3-yl)ethyl-
Scheme 1. Reaction of 4-hydroxy-2H-chromen-2-one 1 with
(N-phenyliminovinylidene)-2a or (2-oxovinylidene)-triphenylpho-
sphorane 2b to 2-oxo-2H-chromen-4-yl-1-N-phenyl-2-(triphenyl-
l5-phosphanylidene) ethanimidoate 3a or 2-oxo-2H-chromen-4-
yl(triphenyl-l5-phosphanylidene) acetate 3b.
phenyl-2H,5H-2l5-[1,2]oxaphosphinino[5,6-c]chromen-5-one 9
was isolated. Compound 9 was obtained through intramo-
lecular cyclization of the intermediate 8 (Scheme 2). The IR
and 1H-NMR spectra of the compound 9 indicated the absence
of the phenolic OH group. Its 31P-NMR spectrum showed
signal at d 21.26 ppm which support the oxaphosphinin
structure [14, 15] and in MS the m/z was found at 462 [M]þ.
X
H
X
PPh3
CH3
O
O
C
O
O
Ph3P=C=C=X
2a, X= N-Ph
b, X= O
CH3
O
O
O
+ Ph3P=O
6a, X= N-Ph
b ,X= O
5
O
O
OH
O
CH3
4
Ph3
P
H
Ph3
P
C=PPh3
O
O
O
O
CH3
CH3
PPh3=C=PPh3
O
7
O
O
9
8
+ Ph3P=O
Scheme 2. Reaction of 3-acetyl-4-hydroxy-2H-chromen-2-one via the complex phosphoranes 5 to give compound 6a and 6b or to react
with the phosphorane 7 to afford compound 9.
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Arch. Pharm. Chem. Life Sci. 2011, 344, 442–450
idene]-4-(triphenyl-l5-phosphanylidene)cyclobutane-1,3-
dione 14b were obtained. Furthermore, a novel approach to
phospholochromenone was performed by interaction of hex-
aphenylcarbodiphosphorane 7 and compound 10 in boiling
tetrahydrofuran. 3-Methyl-(1,1,1-triphenyl-l5-phospholo)[3,2-c]-
chromen-4(1H)-one 15, together with triphenylphosphine
oxide were obtained. The 31P-NMR shift recorded for 15
was d 30.65 ppm and the m/z was found at 446 [M]þ in the
mass spectra (Scheme 3).
phine and triphenylphosphine oxide. The a,b-unsaturated
ketone 16 reacted with the phosphacumulene 2a by
[4 þ 2]-cycloaddition to give the corresponding pyran 17a,
which looses triphenylphospine by Hoffmann degradation
reaction, due to the presence of hydrogen atom in the a-
position to give compound 18. Besides, the phosphacumu-
–
lene 2a undergoes [2 þ 2]-cycloaddtion at the C C of 16 to
–
give the cyclobutylidene 19a, which looses triphenylphos-
phine by Hoffmann degradation to give compound 20.
Moreover, the Wittig reaction between the carbonyl group
of compound 16 and the phosphorane 2a afforded triphenyl-
phosphine oxide and the intermediate ketene, which by
intramolecular cyclization afforded the third product 21a.
The most important feature in the spectroscopic data of the
three products 18, 20, and 21a, is that compound 18 lacks the
presence of carbonyl group in its IR and 13C-NMR and showed
the [M]þ at m/z ¼ 409 in its mass spectra, while compound 20
When 3-[3-(4-fluorophenyl)acryloyl]-2H-chromen-2-one (16)
was treated with (N-phenyliminovinylidene)triphenylphos-
phorane 2a in THF at room temperature for 10 h. Three
products were obtained, namely, 3-[4-(4-fluorophenyl)-2-
(phenylimino)-2H-pyran-6-yl]-2H-chromen-2-one (18), 3-{[2-(4-
fluorophenyl)-4-(phenylimino)cyclobut-2-enecarbon]-1-yl}-2H-
chromen-2-one 20 and 3-(4-fluorostyryl)-1-(phenylimino)cyclo-
penta[c]chromen-4-(1H)-one 21a together with triphenylphos-
Ph3
P
C
X
C
X
O
O
O
C
C
O
Ph3P=C=C=X
CH3
CH3
CH3
2a, X= N-Ph
b, X= O
O
O
O
O
10
12
11
+2
PPh3
X
X
X
CH3
CH3
Ph3P=O
+
O
O
O
O
13a, X= N-Ph
b, X = O
14a, X= N-Ph
b, X= O
Ph3P
CH3
PPh3=C=PPh3
+ Ph3P=O
O
O
7
15
Scheme 3. 3-Acetylchromenone 10 reacts with compound 2a via [2 þ 2] cycloaddition to form oxaphosphetane 11. Elimination of
triphenylphosphine oxide gives 12which undergoes intramolecular cyclization resulting in 13a. Addition of compound 2a,b leads to 13a,b.
The reaction of compound 10 with 7 results in compound 15.
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Arch. Pharm. Chem. Life Sci. 2011, 344, 442–450
New Chromenone Derivatives
445
N
Ph
O
O
H.D
F
+ Ph3P
O
X
18
PPh3
O
F
O
O
17a, X = N-Ph
b, X = O
O
O
H
C
C
F
PPh3=P=C=C=X
H
2a, X = N-Ph
b, X = O
O
16
2a or 2b
O
H
C
H
C
F
O
O
X
PPh3
19a, X = N-Ph
b, X = O
+
X
O
H
H
C
C
C
F
C
H
F
+
O
O
N
Ph
O
O
+ Ph3P=O
+ Ph3P
21a, X = N-Ph
b, X = O
20
Scheme 4. The reaction of compound 16 with phosphacumulene 2b results in compounds 17b, 19b and 21b.
showed the presence of carbonyl group in its IR and 13C-NMR
spectra. Besides, its MS showed the peak at m/z ¼ 409 [M]þ.
The last product lack the presence of the carbonyl group and
in its 1H-NMR a doublet was observed at 5.75 (2JHH ¼ 30 Hz).
Its MS showed the peak at m/z ¼ 393[M]þ.
for 8 h, the corresponding, 3-[4-(4-fluorophenyl)-2-oxo-3-
(triphenyl-l5-phosphanylidene)-3,4-dihydro-2H-pyran-6-yl]-
2H-chromen-2-one 17b, 3-[2-(4-fluorophenyl)ethenyl]cyclopen-
ta[c]chromene-1,4-dione 19b, and 3-{[2-(4-fluorophenyl)-4-oxo-
3-(triphenyl-l5-phosphanylidene)cyclobutyl]carbonyl}-2H-
chromen-2-one 21b accompanied by triphenylphospine oxide
were obtained. The signals at d 21.50 and 29.50 ppm were
obtained in the 31P-NMR spectra of compounds 17b and 19b
respectively (Scheme 4).
The reaction of 3-[3-(4-fluorophenyl)acryloyl]-2H-chromen-
2-one 16 with (2-oxovinylidene)-triphenylphosphorane 2b
was also investigated. When compound 16 was allowed to
react with the phosphacumulene 2b in boiling toluene
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Arch. Pharm. Chem. Life Sci. 2011, 344, 442–450
Figure 1. Doxorubicin (DXR).
Pharmacological evaluation
Chemotherapy is a major approach for both localized and
metastasized cancers [18] and for many years ago coumarin-
related compounds have proved to be a significant thera-
peutic potential [19]. Therefore, four of the newly synthesized
compounds were screened for their in-vitro cytotoxic and
growth inhibitory activities against human breast carcinoma
cell line (MCF-7), in comparison with the activity of the
utilized anticancer doxorubicin (DXR) (Fig. 1) as a reference
drug. The cytotoxic activities of the tested compounds were
expressed as the median growth inhibitory concentration
(IC50) which is the dose that reduces survival to 50%. The
screening results are compiled in Table 1. According to the
American National Cancer Institute guidelines [20] drugs
with IC50 < 30 are active. From Table 1, it is evident that
most of the tested compounds show antitumor activities with
IC50 values ranging from 3.89 to 12.7 mg/mL and reaching
strong correlation that of DXR (IC50: 2.97 mg/mL) in the case
of compound 9 (IC50: 3.89 mg/mL).
Figure 2. The surviving fraction as a function of drug concentration
for the reference compared particularly with the other compounds
(MCF-7).
enzyme topoisomerase II, which unwinds DNA for transcrip-
tion and otherwise stabilizes the topoisomerase II complex
after it has broken the DNA chain for replication, preventing
the DNA double helix from being resealed and thereby stop-
ping the process of replication as act Doxorubicin [21], sup-
pressing agent, inhibit the formation and growth of tumors
from initiated cells [22].
It is clear from the data that the comparison of the cyto-
toxicity against MCF-7 cells (Fig. 2) of prepared compounds
has shown that the cells killing potency follows the order
9 > 6a > 14b > 17b. This may be attributable to presence of
the phospholo-chromenone moiety in its molecular structure
which may contribute to the cytotoxic activity that may
interact with DNA by intercalation and inhibition of macro-
molecular biosynthesis. This inhibits the progression of the
On the other hand, substitution of the phosphanylidene
moiety with the pyran-imine moiety in this compound (cf. 6a,
IC50: 9.99 mg/ml) also closely related the activity of the stan-
dard. The antitumor activity of compound 14b recorded
12.60 mg/ml, this may be due to the presence of phosphor-
anylidene-cyclobutylidene moiety in its molecular structure.
On the other side, compound 17b marked 12.70 mg/mL and
also good result. Therefore, the marked difference in their
activities can be correlated with the difference in biological
activities in different phosphorus and ring systems.
Moreover, the same four compounds (9, 6a, 14b, 17b) were
screened in vitro for their cytotoxic and growth inhibitory
activities towards liver carcinoma (HEPG2) cell line (Fig. 3).
The IC50 after short time exposure was 3.58, 7.04, 9.04, and
14.9 mg/mL for compounds 9, 6a, 14b, and 17b, respectively,
i.e. the cell killing potency follows the order,
9 > 6a > 14b > 17b Table 1. Compound 9 was the best com-
Table 1. Effect of the tested compounds on MCF-7 and HEPG2
tumor cell lines.
Compound No. MCF-7
HEPG2
Doxorubicin (st.)
9
6a
14b
17b
2.97
3.89
9.99
12.60
12.70
3.73
3.58
7.04
9.04
pound, exerting more significant cytotoxicity election (IC50
:
14.90
3.58 mg/mL) HEPG2 cells compared with Doxorubicin (IC50
:
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Arch. Pharm. Chem. Life Sci. 2011, 344, 442–450
New Chromenone Derivatives
447
plentiful source of potential anti-cancer drugs deserving
further study.
Experimental
General
Melting points were determined with an electrothermal digital
melting point apparatus (electrothermal Engineering Ltd., Essex,
UK) and are uncorrected. The IR spectra were recorded in KBr
disks on a Pye Unicam SP 3300 and Shimadzu FT IR 8101 PC
infrared spectrophotometers (Pye Unicam Ltd. Cambridge, UK,
and Shimadzu, Tokyo, Japan, respectively). 1H-, 13C-, and 31P-NMR
spectra were obtained from a Jeol ECA 500 MHz NMR spec-
trometer (Tokyo, Japan) using deuterated dimethylsulphoxide
(d6-DMSO) as a solvent and (TMS) as internal reference at 500, 125,
and 200 MHz, respectively. Mass spectra (EI-MS) were obtained at
70 eV with a Finnigan MAT SSQ 7000 spectrometer (England).
Elemental analyses (C, H, N) results were recorded with
Elementar Vario EL Germany (Germany), P was measured by
spectrophotometric methods and both of them agreed satisfac-
tory with the calculated values. The using reported yields are of
pure isolated materials obtained by column chromatography
silica gel 60 (Merck). Starting materials, 1 [24], 2a [25], 2b [26],
4 [27], 7 [28], 10 [29], and 16 [30] were prepared according to the
procedures reported in the literature.
Figure 3. The surviving fraction as a function of drug concentration
for the reference compared particularly with the other compounds
(HEPG2).
3.73 mg/mL). The results show that compounds 6a and 14b
are better antitumor agents for HEPG2 than MCF-7 and com-
pound 9 may conduct to a promising pharmacomodulation
for the discovery of new potential drugs.
Chemistry
Reaction of (N-phenyliminovinylidene)-2a and (2-
oxovinylidene)-triphenylphosphorane 2b with 4-hydroxy-
2H-chromen-2-one 1
A mixture of 2a (0.01 mol, 0.377 g) or 2b (0.01 mol, 0.302 g) in
20 mL of THF was added drop by drop, with stirring at room
Conclusion
temperature, to
a solution of 4-hydroxychromen-2-one 1
(0.01 mol, 0.162 g) in 20 mL of THF. The reaction mixture was
stirred for 6 h in case of 2a and 10 h in case of 2b during which
the color was changed from white to yellow (the progress of the
reaction was controlled by TLC). The precipitate that formed was
filtered off and crystallized from benzene to give the phosphor-
anylidenes 3a and 3b, respectively.
The nucleophilic phosphacumulene (2a, b) and phosphallene
(7) ylides added reactants in a variety of ways. Cycloaddition
–
occurred at the polar C-P ylide bond or the C C bond.
–
Carbocycles and heterocycles of various ring sizes and het-
eroatom patterns (6, 9, 13, 14, 17, 18, 20, 21), were prepared
which present the pivotal core of many biologically or phar-
maceutically interesting compounds. Moreover, the differ-
ence in the nucleophilic character of the phosphorus
reagents was noticed 2a > 2b > 7 [23]. While the phosphorane
2a reacts smoothly with the reactants, the phosphorane 2b
and 7 react less rapidly.
2-Oxo-2H-chromen-4-yl-1-N-phenyl-2-(triphenyl-l5-
phosphanylidene)ethanimidoate 3a
Colorless crystals, yield 90%, mp: 252–2548C, IR (KBr, n, cmꢀ1
)
1650 (C O, lactone), 1598 (C N), 1579 (C P), 1486, 1471 (P-aryl).
–
–
–
–
–
–
2
1H-NMR (500 MHz, d6-DMSO, d, ppm): 4.5 (d, 1H, JHP ¼ 25 Hz,
CH P) 6.86–7.83 (m, 25H, arom.); 31P-NMR at d ¼ 13.6 ppm. MS
–
–
m/z 538 [M – H]þ. Anal. calcd. for C35H26NO3P (539.5): C, 77.91; H,
4.86; N, 2.60; P, 5.74; Found: C, 77.64; H, 4.55; N, 2.89, P, 5.99.
Many of the new compounds revealed pronounced in-vitro
antitumor activities when tested against human MCF-7 and
HEPG2 carcinoma cell lines. The most promising result
against breast carcinoma (MCF-7) was recorded by the phos-
pholo-chromenone 9. It showed IC50 value of (3.89 mg/mL)
which is the closest in value to that recorded by the reference
drug doxorubicin (IC50: 2.97 mg/mL). Similarly, the cytotoxic
2-Oxo-2H-chromen-4-yl(triphenyl-l5-
phosphanylidene)acetate 3b
Colorless crystals, yield 85%, mp: 231–2338C, IR (KBr, n, cmꢀ1
)
1693 (C O, lactone), 1611 (C O), 1512 (C P), 1484, 1464 (P-aryl).
–
–
–
–
–
–
2
1H-NMR (500 MHz, d6-DMSO, d, ppm): 6.06 (d, 1H, JHP ¼ 22 Hz,
and growth inhibitory activity of the same compound 9 (IC50
:
13
–
–
–
–
CH P) 7.14–7.69 (m, 20H, arom.); C-NMR: 179.63 (C O), 163.27
3.58 mg/mL) was more potent than the same reference drug
(IC50: 3.73 mg/mL) against human liver (HPG2) carcinoma cell
lines. It is concluded that coumarin-related compounds are a
(C O, lactone), 153.64 (C P); 31P-NMR at d ¼ 16.8 ppm. MS m/z:
–
–
–
–
465 [M þ H]þ, 421 [M – CO2]þ. Anal. calcd. for C29H21O4P (464.4):
C, 74.99; H, 4.56; P, 6.67; Found: C, 74.64; H, 4.55, P, 6.09.
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was controlled by TLC). THF was distilled off under reduced
pressure and the residue was subjected to silica gel column
chromatography using petrol ether (60:80)/ethyl acetate as elu-
ent (7:3, v/v) to give 13a and 14a, in addition to triphenylphos-
phine oxide (m.p. and mixed m.p. 1518C).
Reaction of (N-phenyliminovinylidene)-2a or
(2-oxovinylidene)-triphenylphosphorane 2b with 3-acetyl-
4-hydroxy-2H-chromen-2-one 4
A solution of 2a (0.01 mol, 0.377 g) or 2b (0.01 mol, 0.302 g) was
added to solution of 3-acetyl-4-hydroxy-2H-chromene-2-one 4
(0.01 mol, 0.204 g), in toluene (40 mL). The reaction mixture
was boiled for 6 h in case of using 2a and 10 h in case of 2b,
during which the color was changed from colorless to dark brown.
Toluene was distilled off and the residue was subjected to silica gel
column chromatography using petrol ether (60:80)/ethyl acetate
as eluent (65:35, v/v) to give 6a and 6b respectively together with
triphenylphosphine oxide (m.p. and mixed m.p. 1518C).
3-Methyl-1-(phenylimino)cyclopenta[c]chromen-4(1H)-
one 13a
Orange crystals, yield 55%, mp: 333–3358C. IR (KBr, n, cmꢀ1) 1726
1
(C O, lactone), 1618 (C N). H-NMR (500 MHz, d -DMSO, d, ppm):
–
–
–
–
6
2.16 (s, 3H, CH3), 4.75 (s, 1H, CH), 7.22–7.30 (m, 9H, arom.): MS m/z:
286 [M – H]þ; 241 [M – CO2]þ. Anal. calcd. for C19H13NO2 (287.3): C,
79.43; H, 4.56; N, 4.88; Found: C, 79.16; H, 4.44; N, 4.13.
4-Methyl-2-(phenylimino)pyrano[3,2-c]chromen-5(2H)-
one 6a
3-{1-[2,4-Bis(phenylimino)-3-(triphenyl-l5-
phosphanylidene)cyclobutylidene]ethyl}-2H-chromen-2-
one 14a
Colorless crystals, yield 80%, mp: 210–2128C, IR (KBr, n, cmꢀ1
)
1712 (C O, lactone), 1608 (C N). H-NMR (500 MHz, d -DMSO, d,
1
–
–
–
ppm): 2.19 (s, 3H, CH3), 5.89 (s, 1H, CH), 6.97–7.92 (m, 9H, arom.).
–
6
Yellow crystals, yield 40%, mp: 178–1808C, IR (KBr, n, cmꢀ1) 1729
13
–
–
–
C-NMR: 163.58 (C N), 162.18 (C O, lactone), 25.92 (CH ). MS m/z:
3
– –
(C O, lactone), 1682, 1598 (2C N), 1563 (C P), 1490, 1458 (P-aryl).
– –
–
–
–
304 [M þ H]þ. Anal. calcd. for C19H13NO3 (303.31): C, 75.24; H,
1H-NMR (500 MHz, d6-DMSO, d, ppm): 2.72 (s, 3H, CH3), 7.30–7.85
13
(m, 30H, arom.). C-NMR: 163.03 (2 C N), 159.89 (C O, lactone),
4.32; N, 4.62; Found: C, 75.70; H, 4.44; N, 4.33.
–
–
–
–
153.75 (C P), 22.12 (CH ). 31P-NMR d ¼ 30.03 ppm. MS m/z: 663 [M –
–
–
3
H]þ, 648 [M – CH3]þ. Anal. calcd. for C45H33N2O2P (664.7): C, 81.31;
H, 5.00; N, 4.21; P, 6.44; Found: C, 80.92; H, 4.88; N, 4.05; P, 6.28.
4-Methyl-2H, 5H-pyrano-[3,2-c]chromene-2,5-dione 6b
Pale yellow crystals, yield 35%, mp: 147–1498C. IR (KBr, n, cmꢀ1
)
1704 (C O, lactone), 1671 (C O). H-NMR (500 MHz, d -DMSO, d,
1
–
–
–
ppm): 2.30 (s, 3H, CH3), 6.36 (s, 1H, CH), 7.25–7.66 (m, 4H, arom.).
–
6
Reaction of (2-oxovinylidene)triphenylphosphorane 2b
with 3-acetyl-2H-chromen-2-one 10
13
–
–
–
C-NMR: 162.00 (C O, lactone), 161.14 (C O, lactone), 22.21 (CH ).
3
–
MS m/z: 228 [M]þ, 200 [M – CO]þ, 184 [M – CO2]þ. Anal. calcd.
for C13H8O4 (228.2): C, 68.42; H, 3.53; Found: C, 68.02; H, 3.29.
A mixture of 2b (0.01 mol, 0.302 g), 10 (0.01 mol, 0.188 g), and
toluene (40 mL) was refluxed for 8 h during which the color
changed from yellow to dark brown. Toluene was distilled off
and the residue was subjected to silica gel column chromatog-
raphy using petrol ether (60:80)/ethyl acetate as eluent (3:7, v/v)
to give 13b and 14b in addition to triphenylphosphine oxide
(m.p. and mixed m.p. 1518C).
Interaction of hexaphenylcarbodiphosphorane 7 with
3-acetyl-4-hydroxy-2H-chromen-2-one 4
To a solution of 4 (0.01 mol, 0.204 g) in 20 mL THF was added 7
(0.01 mol, 0.536 g) in 20 mL THF. The reaction mixture was
refluxed for 12 h during which the color change from colorless
to yellow then brown. THF was distilled off under reduced
pressure and the remained residue was chromatographed on
silica gel using petrol ether (60:80)/ethyl acetate as eluent (60:40,
v/v) to give 9 together with triphenylphosphine oxide (m.p. and
mixed m.p. 1518C).
3-Methylcyclopenta[c]chromene-1,4-dione 13b
Colorless crystals, yield 49%, mp: 110–1128C. IR, (KBr, n, cmꢀ1
)
1727 (C O, lactone), 1603 (C O). H-NMR (500 MHz, d -DMSO, d,
1
–
–
–
ppm): 2.21 (s, 3H, CH3), 5.93 (s, 1H, CH), 7.26–7.56 (m, 4H, arom.).
–
6
13
–
–
–
C-NMR: 180.00 (C O), 159.70 (C O, lactone), 22.06 (CH ). MS
3
–
m/z: 213 [M þ H]þ, 211 [M – H]þ. Anal. calcd. for C13H8O3 (212.20):
4-Methyl-2,2,2-triphenyl-2H,5H-2l5-
[1,2]oxaphosphinino[5,6-c]chromen-5-one 9
C, 73.58; H, 3.80; Found: C, 72.99; H, 3.76.
2-[1-(2-Oxo-2H-chromen-3-yl)ethylidene]-4-(triphenyl-l5-
phosphanylidene)cyclobutane-1,3-dione 14b
Colorless crystals, yield 75%, mp: 233–2358C. IR (KBr, n, cmꢀ1) 1690
1
(C O, lactone), 1561 (C P), 1480, 1434 (P-aryl). H-NMR (500 MHz,
–
–
–
–
d6-DMSO, d, ppm): 1.94 (s, 3H, CH3), 7.25–7.58 (m, 20H, arom.).
Pale yellow crystals, yield 45%, mp: 263–2658C. IR, (KBr, n, cmꢀ1
)
1722 (C O, lactone), 1633, 1620 (2 C O), 1590 (C P), 1482, 1436
C-NMR: 177 (C O), 161.14 (C O, lactone), 24.71 (CH ). 31P-NMR:
13
–
–
–
–
–
–
–
3
–
–
–
21.26. MS m/z: 462 [M]þ, 463 [M þ H]þ. Anal. calcd.
for C30H23O3P (462.4): C, 77.91; H, 5.01; P, 6.70; Found: C, 77.02;
H, 4.99; P, 6.06.
(P-aryl). 1H-NMR (500 MHz, d6-DMSO, d, ppm): 2.52 (s, 3H, CH3),
7.55–7.71 (m, 20H, arom.). 13C-NMR: 186.29 (2 C O), 159.90 (C O,
–
–
–
–
lactone), 153.66 (C P), 16.67 (CH ). 31P-NMR d ¼ 29.79 ppm.
–
–
3
MS m/z: 515 [M þ H]þ, 514 [M]þ, 513 [M – H]þ, 486 [M – CO]þ,
469 [M – CO2]þ, 457 [M – 2 CO]þ. Anal. calcd. for C33H23O4P (514.2):
C, 77.04; H, 4.51; P, 6.02; Found: C, 76.92; H, 4.06; P, 6.03.
Reaction of (N-phenyliminovinylidene)
triphenylphosphorane 2a with 3-acetyl-2H-chromen-
2-one 10.
A solution of 2a (0.377 g, 0.01 mol) in 20 mL of THF was added
drop by drop, with stirring at room temperature, to a solution of
10 (0.01 mol, 0.188 g) in 20 mL of THF for 10 h, during which the
color changed from yellow to brown (the progress of the reaction
Interaction of hexaphenylcarbodiphosphorane 7 with
3-acetyl-2H-chromen-2-one 10
To a solution of 10 (0.01 mol, 0.188 g) in 20 mL THF was added 7
(0.01 mol, 0.536 g) in 20 mL THF. The reaction mixture was
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Arch. Pharm. Chem. Life Sci. 2011, 344, 442–450
New Chromenone Derivatives
449
refluxed for 12 h during which the color change from colorless
to yellow then brown. THF was distilled off under reduced
pressure and the remained residue was chromatographed on
silica gel using petrol ether (60:80)/ethyl acetate as eluent (60:40,
v/v) to give 15 and together with triphenylphosphine oxide (m.p.
and mixed m.p. 1518C).
Interaction of (2-oxovinylidene)triphenylphosphorane 2b
and 3-(3-(4-fluorophenyl)acryloyl)-2H-chromen-2-one 16
A mixture of 2b (0.01 mol, 0.302 g), 16 (0.01 mol, 0.294 g), and
toluene (40 mL) was refluxed for 8 h during which the color
changed from yellow to dark brown. Toluene was distilled off
and the residue was subjected to silica gel column chromatog-
raphy using petrol ether (60:80)/ethyl acetate as eluent (6:4, v/v),
to give 17b, 19b, and 21b in addition to triphenylphosphine
oxide (m.p. and mixed m.p. 1518C).
3-Methyl-(1,1,1-triphenyl-l5-phospholo)[3,2-c]chromen-
4(1H)-one 15
Colorless crystals, yield 70%, mp: 103–1058C. IR, (KBr, n, cmꢀ1
)
–
–
3-[4-(4-Fluorophenyl)-2-oxo-3-(triphenyl-l5-
phosphanylidene)-3,4-dihydro-2H-pyran-6-yl]-2H-
1715 (C O, lactone), 1596 (C P), 1481, 1432 (P-aryl). 1H-NMR
–
–
(500 MHz, d6-DMSO, d, ppm): 2.02 (s, 3H, CH3), 7.24–7.73 (m,
20H, arom.). 31P-NMR: 30.65. MS m/z: 446 [H]þ, 444 [M – 2 H]þ.
Anal. calcd. for C30H23O2P (446.4): C, 80.70; H, 5.19; P, 6.94;
Found: C, 80.14; H, 5.06; P, 6.20.
chromen-2-one 17b
Yellow crystals, yield 40%, mp: 247–2498C. IR (KBr, n, cmꢀ1) 1716
–
–
–
(C O, lactone), 1632 (C O, lactone), 1502 (C P), 1434, 1356 (P-
–
–
–
aryl). 1H-NMR (500 MHz, d6-DMSO, d, ppm): 6.72–7.55 (m, 25H,
arom.); 8.44 (s, 1H, CH, methine); 13C-NMR: 165.82 (C O, lactone),
–
–
Reaction of (N-phenyliminovinylidene)
triphenylphosphorane 2a with 3-(3-(4-fluorophenyl)
acryloyl)-2H-chromen-2-one 16
160.24 (C O, lactone), 152.79 (C P). 31P-NMR: d ¼ 21.50 ppm. MS
–
–
–
–
m/z: 598 [M þ 2 H]þ, 596 [M]þ. Anal. calcd. for C38H26FO4P (596.5):
C, 75.50; H, 4.39; F, 3.18; P, 5.19; Found: C, 75.02; H, 4.29; F, 3.04;
P, 5.11.
A solution of 2a (0.377 g, 0.01 mol) in 20 mL of THF was added
drop by drop, with stirring at room temperature, to a solution of
16 (0.01 mol, 0.294 g) in 20 mL of THF for 10 h, during which the
color changed from yellow to brown (the progress of the reaction
was controlled by TLC). THF was distilled off under reduced
pressure and the residue was subjected to silica gel column
chromatography using petrol ether (60:80)/ethyl acetate as elu-
ent (7:3, v/v), to give 18, 20, and 21a in addition to triphenyl-
phosphine (m.p. and mixed m.p. 788C) and triphenylphosphine
oxide (m.p. and mixed m.p. 1518C).
3-{[2-(4-Fluorophenyl)-4-oxo-3-(triphenyl-l5-
phosphanylidene)cyclobutyl]carbonyl}-2H-chromen-2-one
19b
Yellow crystals, yield 35%, mp: 147–1498C. IR (KBr, n, cmꢀ1) 1715
(C O, lactone), 1620 (C O), 1599 (C P), 1437, 1300 (P-aryl). 1H-
NMR (500 MHz, d6-DMSO, d, ppm): 2.21 (m, 1H, CH); 3.03 (d, 1H,
–
–
–
–
–
–
2JHH ¼ 15 Hz, CH), 6.28–7.66 (m, 24H, arom.); 13C-NMR: 200.10
(C O), 159.24 (C O, lactone), 152.79 (C P). 31P-NMR:
–
–
–
–
–
–
d ¼ 29.50 ppm. MS m/z: 597 [M þ H]þ, 596 [M]þ. Anal. calcd.
for C38H26FO4P (596.5): C, 75.50; H, 4.39; F, 3.18; P, 5.19;
Found: C, 75.02; H, 4.19; F, 3.04; P, 4.99.
3-[4-(4-Fluorophenyl)-2-(phenylimino)-2H-pyran-6-yl]-2H-
chromen-2-one 18
Yellow crystals, yield 35%, mp: 245–2478C. IR (KBr, n, cmꢀ1) 1715
1
(C O, lactone), 1622 (C N). H-NMR (500 MHz, d -DMSO, d, ppm):
–
–
–
–
6
3-[2-(4-Fluorophenyl)ethenyl]cyclopenta[c]chromene-1,4-
dione 21b
7.08–7.50 (m, 15H, arom.); 8.65 (s, 1H, CH, methine); 13C-NMR:
–
–
168.65 (C N), 162.54 (C-F), 161.36 (C O, lactone). MS m/z: 409
–
–
Yellow crystals, yield 25%, mp: 185–1878C. IR (KBr, n, cmꢀ1) 1717
[M]þ, 408 [M – H]þ, 380 [M – CO]þ. Anal. calcd. for C26H16FNO3
(409.4): C, 76.42; H, 3.94; F, 4.64; N, 3.42; Found: C, 76.02; H, 3.49;
F, 4.04; N, 3.22.
1
(C O, lactone), 1633 (C O). H-NMR (500 MHz, d -DMSO, d, ppm):
–
–
–
–
6
6.44 (d, 1H, JHH ¼ 15 Hz, CH), 7.49–769 (m, 10H, arom.); 13C-
2
þ
–
–
–
–
NMR: 187.44 (C O), 161.70 (C O, lactone). MS m/z: 275 [M – CO ] .
2
Anal. calcd. for C20H11FO3 (318.3): C, 75.47; H, 3.48; F, 5.97;
Found: C, 75.02; H, 3.40; F, 5.24.
3-{[2-(4-Fluorophenyl)-4-(phenylimino)cyclobut-2-
enecarbon]-1-yl}-2H-chromen-2-one 20
Orange crystals, yield 30%, mp: 237–2398C. IR (KBr, n, cmꢀ1) 1729 Material and methods
(C O, lactone), 1671 (C O), 1603 (C N). 1H-NMR (500 MHz, d6-
DMSO, d, ppm): 4.17 (s, 1H, CH), 6.07 (s, 1H, CH), 7.10–7.85 (m,
–
–
–
–
–
–
Chemicals
13
13H, arom.); 8.28 (s, 1H, CH, methine); C-NMR: 186.28 (C O),
–
–
163.27 (C N), 159.98 (C–F), 159.33 (C O, lactone). MS m/z: 409
All the chemicals and reagents used in this study were of
analytical grade and purchased from (Sigma Chemical Co., St.
Louis, MO, USA): These were used in cryopreservation of cells.
–
–
–
–
[M]þ. Anal. calcd. for C26H16FNO3 (409.4): C, 76.42; H, 3.94; F, 4.64;
N, 3.42; Found: C, 76.02; H, 3.49; F, 4.04; N, 3.22.
3-(4-Fluorostyryl)-1-(phenylimino)cyclopenta[c]chromen-
4(1H)-one 21a
Cells culture
The cells of MCF-7 human breast cancer and HEPG2 liver
carcinoma were maintained and grown in RPMI-1640
medium supplemented with 10% heat inactivated fetal
bovine serum (Sigma Chemical Co., St. Louis, MO, USA),
penicillin and streptomycin at 378C in humidified atmo-
sphere containing 5% CO2.
Yellow crystals, yield 30%, mp: 147–1498C. IR (KBr, n, cmꢀ1) 1718
1
–
–
–
–
(C O, lactone), 1586 (C N). H-NMR (500 MHz, d -DMSO, d, ppm):
6
5.76 (d, 1H, 2JHH ¼ 30 Hz, CH), 5.83 (s, 1H, CH), 7.01–7.71 (m, 14H,
arom.). MS m/z: 393 [M]þ. Anal. calcd. for C26H16FNO2 (393.4): C,
79.38; H, 4.10; F, 3.83; N, 3.56; Found: C, 79.12; H, 3.99; F, 3.64; N,
3.22.
ß 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com

450
S. S. Maigali et al.
Arch. Pharm. Chem. Life Sci. 2011, 344, 442–450
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Article ID 159925. DOI: 10.1155/2007/15925.
In-Vitro cytotoxicity assay
For in-vitro short term cytotoxicity evaluation of prepared
compounds, MCF-7 and HEPG2 cells were plated a concen-
tration of 5 ꢁ 104–105 cells per well, in complete culture
medium in 96-well flat-bottomed culture plates (Falcon) for
24 h to assure total attachment. Then various concentration
of test compounds were added to the cell suspended in
0.10 ml of phosphate buffered saline (FBS) (0.20 M, pH 7.4),
the control cells without the test compounds were also cul-
tured, then the plate was incubated for 24 h at 408C and 72 h
at 378C, in a humidified 5% CO2 atmosphere. Cell survival was
evaluated at the end of the incubation period with sulpho-
rhodamine-B (SRB) colorimetric assay according to Skehan
et al. [20]. This test is based on the sensitivity of the human
tumor cell lines to thymoquinone was determined by the SRB
assay. SRB is a bright pink aminoxanthrene dye with two
sulphonic groups. It is a protein stains that binds to the
amino groups of intracellular proteins under mildly acidic
conditions to provide a sensitive index of cellular protein
content. After incubation, media were removed and 50 mL of
0.4% SRB dissolved in 1% acetic acid solution well were. The
wells were then washed 4 times with 1% acetic acid. The
absorbance was determined photometrically at 564 nm with
ELISA microplate reader (Meter tech. S 960, USA.).
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The percentage of cell survival was calculated as follows:
Survival fraction ¼ O.D. (treated cells)/O.D. (control cells)
where (O. D.) is the optical density. The IC50 values (the
concentrations of thymoquinone required to produce 50%
inhibition of cell growth). The experiment was repeated 3
times for each cell line.
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´
´
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We are grateful to the Cancer Biology Department, National Cancer
Institute, Cairo University for the pharmacological evaluation.
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The authors have declared no conflict of interest.
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ß 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com