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SYNTHESIS, REACTIONS OF 3-OXO-3-(3-OXO-3H-BENZO[f] CHROMEN-2-

YL)-2-(2-PHENYLHYDRAZONO) PROPANAL, AND INVESTIGATION OF THEIR
ANTITUMOR ACTIVITY

Article  in  WORLD JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES · August 2013

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 WORLD JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES
Sobhi et al. World Journal of Pharmacy and Pharmaceutical Sciences
Volume 2, Issue 5, 2341-2354. Research Article ISSN 2278 – 4357

SYNTHESIS, REACTIONS OF 3-OXO-3-(3-OXO-3H-BENZO[f]

CHROMEN-2-YL)-2-(2-PHENYLHYDRAZONO) PROPANAL, AND
INVESTIGATION OF THEIR ANTITUMOR ACTIVITY

Dr Sobhi M. Gomhaa,* and Hassan M. Abdel-aziz b

a
Department of Chemistry, Faculty of Science, University of Cairo, Giza, 12613, Egypt.
b
Department of Chemistry, Faculty of Science, Bani Suef University, Bani Suef, Egypt.

Article Received on ABSTRACT

01 August 2013,
Heteroarylhydrazonalas were prepared from coupling of sodium salt of
Revised on 29 August 2013,
2-(3-hydroxyacryloyl)-3H-benzo[f]chromen-3-one with various
Accepted on 25 September
2013 diazotized heterocyclic amines. Treatment of heteroarylhydrazonalas,
with 2-cyanoacetamide, 3-oxo-3-phenylpropanenitrile,
*Correspondence for ethylcyanoactate, and -haloketones afforded the corresponding
Author:
pyridazinone, pyridine, and pyrazole derivatives, respectively. The
* Dr Sobhi M. Gomha synthesized compounds were characterized on the basis of their
Department of Chemistry, elemental analysis and spectral data. All the newly synthesized
Faculty of Science, University
compounds were evaluated for their antitumor activities against the
of Cairo, Giza, 12613, Egypt.
human breast cancer cell line MCF-7 and the liver carcinoma cell line
s.m.gomha@hotmail.com
HEPG-2, and the results of some derivatives showed promising
activity.
Keywords: Coupling reaction, pyrazolotriazine, triazolotriazine, benzoimidazotriazine
pyridazinone, and antitumor evaluation.

INTRODUCTION
Coumarins are a class of compounds with wide and varied biological activity. They are
known to be analgesics,1 anticancer,2,3 and antibacterial agents4,5 as well as being specific
inhibitors of α-chymotripsin,6 human leukocyte elastase,7 platelet aggregation,8,9 inhibitor of
10
HIV-1 protease. Moreover, coumarin-based dyes and pigments are organic fluorescent
materials exhibiting unique photochemical and photophysical properties, which render them

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 Sobhi et al. World Journal of Pharmacy and Pharmaceutical Sciences

useful in a variety of applications such as dye lasers, anion sensors, organic light-emitting
diodes and solar cells. 11, 12
In view of the above mentioned findings and as continuation of our efforts in the synthesis of
new biologically active heterocycles,13-21 we report herein the results of our study of the
coupling of sodium salt of 2-(3-hydroxyacryloyl)-3H-benzo[f]chromen-3-one with several
diazotized heterocyclic amines.

RESULTS AND DISCUSSION

In the first instance, sodium salt of 2-(3-hydroxyacryloyl)-3H-benzo[f]chromen-3-one (2)
was synthesized by reported method 21 from 2-acetyl-3H-benzo[f]chromen-3-one 1 with ethyl
formate in dry ether containing sodium methoxide.

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Further, the reactivity of salt 2 towards some diazotized heterocyclic amines was examined.
Thus, coupling of 2 with diazotized aminopyridine 3, diazotized aminopyrimidine 4,
diazotized aminothiazole 5, diazotized aminobenzothiazole 6, diazotized aminoantipyrene 7
in ethanol containing sodium acetate yielded the respective heteroarylhydrazonalas
1
derivatives 8-12 (Scheme 1). The H NMR spectrum of each of the products 8-12 revealed
two singlet signals in the regions δ 10.13-10.18(D2O exchanged) and 13.36-13.57 assignable
to the hydrazone-NH proton and CHO proton, respectively. Similar treatment of 2 with
diazonium salts of 3-phenyl-1H-pyrazol-5-amine 13, 4H-1,2,4-triazol-3-amine 14, and 2-
aminobenzimidazole 15 under the same reaction conditions afforded the respective
pyrazolo[5,1-c][1,2,4]triazine 16, triazolo[3,4-c][1,2,4]triazine 17, and benzoimidazo [2,1-
c][1,2,4]triazine 18 derivatives, respectively (Scheme 2). The structure of compounds 16, 17,
and 18 were elucidated on the basis of spectroscopic data and microanalysis as expected (See
experimental section). The 1H NMR spectrum of compound 16, taken as a typical example
of the series, exhibited a revealed signal at δ 9.67 due to a proton of triazine-H5, in addition
to a multiplet at δ 6.73-8.11 due to 12 aromatic protons assignable to phenyl, pyrazole, and
naphthocoumarine protons. Also, its mass spectrum revealed molecular ion peak at m/z 418.

The formation of products 16-18 seems to result via initial coupling of diazonium salt on
active methylene group in compound 2 to form the respective non-isolable azo coupling
21-23
intermediate A rather than B, which then underwent intramolecular dehydrative
cyclization to give the isolated products 16-18 (cf. Scheme 2). In addition,
heteroarylhydrazonalas derivative 12 reacted with 2-cyanoacetamide 19a, 3-oxo-3-
phenylpropanenitrile 19b, and ethylcyanoactate 19c in acetic acid containing ammonium
acetate under reflux, afforded the corresponding pyridazinone 21a,b and pyridine derivatives
22,24-26 respectively(Scheme 3). The formation of pyridazinone derivatives 21a,b is suggested
to proceed through the initial formation of imino derivatives 20a,b, followed by readily
hydrolysis of imino group into oxo group giving 21a,b. The structure of the isolated product
was elaborated by its elemental analyses and spectral data (IR, 1H NMR, and MS) (see
Experimental). IR spectrum of 21a showed absorption bands at v = 3189, 3358 cm-1
assignable to NH2 group besides carbonyl absorption bands at v = 1644 -1727 cm−1. Also, its
1
H NMR spectrum revealed singlet signal at δ 11.34 ppm (D2O-exchangeable) due to NH2
protons.

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 Sobhi et al. World Journal of Pharmacy and Pharmaceutical Sciences

27
Finally, similar to recent report, the reaction of 12 with chloroacetone 23a and phenacyl
bromide 23b in ethanol containing K2CO3 at room temperature afforded the corresponding

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pyrazole derivatives 24a,b, respectively (Scheme 4). The structures of the products 24a,b
1
were consistent with their elemental and spectral analyses. Thus H NMR spectrum of 24a
showed 3 singlet signals at δ = 2.13, 2.37 and 2.48 ppm assignable for CH3CO, CH3 and
NCH3, respectively. in addition to multiplet bands at 6.57-8.18 ppm for aromatic protons.
Moreover, the mass spectrum give an additional conformation for the correct structure which
showed the molecular ion peak m/z = 518 (M+ , 33%) (Scheme 4).

Pharmacology
Anticancer activity
The in vitro antitumor activity of the tested compounds was evaluated at Regional Center for
Mycology and Biotechnology at Al-Azhar University, Cairo, Egypt. Thirteen compounds
were tested for their in vitro antitumor activity against the HEPG-2 ‘‘liver’’cancer cell line
and the MCF-7 ‘‘breast’’ cancer cell line. Doxorubicin was used as a reference and showed
IC50 = 0.482 and 0.436 g/mL against the HEPG-2 and MCF-7 cell lines, respectively. Some
of the tested compounds showed effects against both cancer cell lines as shown in Table 3.
Considering the observed antitumor screening data against the MCF7 ‘‘breast’’ cancer cell
line, only compounds 12, 17, 18, 21a, 22, and 24b revealed promising pharmacological
activity (IC50 = 0.86, 0.51, 1.82, 1.23, 6.2, and 0.54 g/mL, respectively) compared with
doxorubicin which was used as a reference standard during this study (IC50 = 0.436 g/mL)
(Table 1).
Also, compounds 12, 17, 18, 21a, 21b, 24a, and 24b exhibited good activity against the
HEPG2 ‘‘liver’’ cell line (IC50 = 0.72, 0.62, 2.8, 1.04, 2.54, 0.64, and 1.46 g/mL,
respectively) compared with doxorubicin (IC50 = 0.482 g/mL).

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Structure–activity relationship based on the observed antitumor properties of the synthesized

compounds against both of the two cancer cell lines indicated that attachment of
naphthocoumarine ring with antipyrine ring seemed more favorable for exhibiting antitumor
properties.

Table 1. Cytotoxic activity of the synthesized compounds against the HEPG-2 and
MCF-7 cell lines.
Compd. no. IC50 (g/mL) IC50 (g/mL)
HEPG-2 cell line MCF-7 cell line
8 20.4 21.7
9 17.9 15.8
10 212 21.4
11 32.5 31.1
12 0.72 0.86
16 33.3 22.3
17 0.62 0.51
18 2.80 1.82
21a 1.04 1.23
21b 2.54 56.43
22 14.84 6.2
24a 0.64 10.5
24b 1.46 0.54
Doxorubicin 0.482 0.436

EXPERIMENTAL
All melting points were measured on Electro thermal IA 9000 series digital melting point
apparatus. The IR spectra were recorded in potassium bromide discs on a Pye Unicam SP
3300 and Shimadzu FT IR 8101 PC infrared spectrophotometer. The NMR Spectra were
recorded at 270 MHz on a Varian Mercury VX-300 NMR spectrometer. 1H NMR (300 MHz)
was run in deuterated dimethylsulphoxide (DMSO-d6). Chemical shifts were related to that of
the solvent. Mass Spectra were recorded on a Shimadzu GCMS-QP1000 EX mass
spectrometer at 70 eV. Elemental analyses and the antitumor evaluation of the products were
carried out at the Microanalytical Center of Cairo University, Giza, Egypt. All reactions were

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followed by TLC (Silica gel, Aluminum Sheets 60 F254, Merck). Sodium salt of 2-(3-
hydroxyacryloyl)-3H-benzo[f]chromen-3-one 220 was prepared as reported in the literature.

Coupling of salt 2 with diazotized heterocyclic amines

General procedure. To a solution of sodium salt of 2-(3-hydroxyacryloyl)-3H-
benzo[f]chromen-3-one 2, (0.5 g, 2 mmol) in EtOH (20 mL) was added sodium acetate
o
trihydrate (0.276 g, 2 mmol), and the mixture was cooled to 0-5 C in an ice bath. To the
resulting cold solution was added portionwise a cold solution of the appropriate diazotized
heterocyclic amines, namely, 2-aminopyridine 3, 2-aminopyrimidine 4, 2-aminothiazole 5, 2-
aminobenzothiazole 6, 2-aminoantipyrene 7, 3-phenyl-1H-pyrazol-5-amine 13, 4H-1,2,4-
triazol-3-amine 14, and 2-aminobenzimidazole 15 [prepared by diazotizing heterocyclic
amine derivatives, (2 mmol) dissolved in hydrochloric acid (6 M, 2 mL) or in nitric acid
(6M, 2ml) or in sulphuric acid (6M, 1ml) with a solution of sodium nitrite (0.14 g, 2 mmol)
in water (5 mL)]. After complete addition of the diazonium salt, the reaction mixture was
stirred for a further 30 min in an ice bath. The reaction mixture was then left in a refrigerator
for two days. The solid that precipitated was filtered off, washed with water, dried and finally
crystallized from ethanol to give the respective products 8-12 and 16-18.

3-Oxo-3-(3-oxo-3H-benzo[f]chromen-2-yl)-2-(2-(pyridin-2-yl)hydrazono)propanal (8):
Yellow solid (81%); m.p. 226-8 °C (EtOH); IR: v 1618(C=N), 1696, 1720, 1742(3C=O),
3218(NH) cm-1 ; 1H NMR: δ 6.86-8.04(9H, m, Ar-H), 8.12(1H, s, pyrimidine-H4), 9.18(1H, s,
Coumarin-H4), 10.18(1H, s, br, NH), 13.57(1H, s, CHO); MS m/z (%): 372(M+ + 1, 11),
371(M+, 82), 239(32), 193(34), 151(56), 84(100). Anal. Calcd for C21H13N3O4 (371.35): C,
67.92; H, 3.53; N, 11.32. Found C, 67.76; H, 3.43; N, 11.24%.

3-Oxo-3-(3-oxo-3H-benzo[f]chromen-2-yl)-2-(2-(pyrimidin-2-yl)hydrazono)propanal
(9): Yellow solid (76%); m.p. 248-9 °C (EtOH); IR: v 1618(C=N), 1688, 1724, 1740(3C=O),
3233(NH) cm-1 ; 1H NMR: δ 6.88-8.48(9H, m, Ar-H), 9.23(1H, s, Coumarin-H4), 10.13(1H,
s, br, NH), 13.36(1H, s, CHO); MS m/z (%): 373(M+ + 1, 8), 372(M+, 34), 195(90), 168(37),
84(69), 63(100). Anal. Calcd for C20H12N4O4 (372.33): C, 64.52; H, 3.25; N, 15.05. Found C,
64.43; H, 3.20; N, 15.00%.

3-Oxo-3-(3-oxo-3H-benzo[f]chromen-2-yl)-2-(2-(thiazol-2-yl)hydrazono)propanal (10):
Yellow solid (79%); m.p. 207-9 °C (EtOH); IR: v 1618(C=N), 1696, 1720, 1745(3C=O),

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3233(NH) cm-1 ; 1H NMR: δ 6.82-8.11(8H, m, Ar-H), 9.22(1H, s, Coumarin-H4), 10.16(1H,

s, br, NH), 13.50(1H, s, CHO); MS m/z (%): 378(M+ + 1, 12), 377(M+, 100), 319(32),
130(27), 77(54). Anal. Calcd for C19H11N3O4S(377.37): C, 60.47; H, 2.94; N, 11.13. Found
C, 60.28; H, 2.76; N, 11.01%.

2-(2-(Benzo[d]thiazol-2-yl)hydrazono)-3-oxo-3-(3-oxo-3H-benzo[f]chromen-2-
yl)propanal
(11): Yellow solid (77%); m.p. 186-8 °C (EtOH); IR: v 1631(C=N), 1690, 1734,
1741(3C=O), 3247(NH) cm-1; 1H NMR: δ 6.82-8.11(10H, m, Ar-H), 9.26(1H, s, Coumarin-
H4), 10.10(1H, s, br, NH), 13.47(1H, s, CHO); MS m/z (%): 428(M+ + 1, 9), 427(M+, 9),
194(54), 144(100), 51(49). Anal. Calcd for C23H13N3O4S (427.43): C, 64.63; H, 3.07; N,
9.83. Found C, 64.76; H, 3.08; N, 9.77%.

2-(2-(1,5-Dimethyl-3-oxo-2-phenyl-2,3-dihydro-1H-pyrazol-4-yl)hydrazono)-3-oxo-3-(3-
oxo-3H-benzo[f]chromen-2-yl)propanal (12): Yellow solid (78%); m.p. 196-8 °C (EtOH);
IR: v 1631(C=N), 1669, 1692, 1717, 1748(4C=O), 3211(NH) cm-1 ; 1H NMR: δ 2.51(3H, s,
CH3), 3.48(1H, s, NCH3), 6.65-8.08(11H, m, Ar-H), 9.22(1H, s, Coumarin-H4), 10.13(1H, s,
br, NH), 13.43(1H, s, CHO); MS m/z (%): 481(M+ + 1, 14), 480(M+, 100), 239(34), 193(26),
151(38), 84(65). Anal. Calcd for C27H20N4O5 (480.47): C, 67.49; H, 4.20; N, 11.66. Found C,
67.42; H, 4.29; N, 11.43%.

2-(7-Phenylpyrazolo[5,1-c][1,2,4]triazine-3-carbonyl)-3H-benzo[f]chromen-3-one (16):
Yellow solid (71%); m.p. 276 °C (DMF); IR: v 1683, 1725(2C=O) cm-1 ; 1H NMR: δ 6.73-
8.11(12H, m, Ar-H), 9.27(1H, s, Coumarin-H4), 9.67(1H, s, triazine-H5); MS m/z (%):
419(M+ + 1, 24), 418(M+, 41), 263(35), 194(36), 144(100), 84(59), 55(54). Anal. Calcd for
C25H14N4O3(418.40): C, 71.77; H, 3.37; N, 13.39. Found C, 71.58; H, 3.39; N, 13.19%.

2-([1,2,4]Triazolo[3,4-c][1,2,4]triazine-6-carbonyl)-3H-benzo[f]chromen-3-one (17):
Yellow solid (75%); m.p. 249 °C (DMF); IR: v 1617(C=N), 1692, 1730(2C=O) cm-1 ; 1H
NMR: δ 6.73-8.11(6H, m, Ar-H), 8.69(1H, s, triazole-H5), 9.22(1H, s, Coumarin-H4),
9.67(1H, s, triazine-H5); MS m/z (%): 344(M+, 4), 343(91), 304(21), 178(13), 105(100),
77(92). Anal. Calcd for C18H9N5O3 (343.30): C, 62.98; H, 2.64; N, 20.40. Found C, 62.78; H,
2.54; N, 20.27%.
2-(Benzo[4,5]imidazo[2,1-c][1,2,4]triazine-3-carbonyl)-3H-benzo[f]chromen-3-one (18):
Yellow solid (77%); m.p. 277-9 °C (DMF); IR: v 1617(C=N), 1678, 1716(2C=O) cm-1; 1H

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NMR: δ 6.73-8.11(10H, m, Ar-H), 9.24(1H, s, Coumarin-H4), 9.56(1H, s, triazine-H5); MS

m/z (%): 392 (M+, 8), 197 (19), 84 (100), 56 (48). Anal. Calcd for C23H12N4O3 (392.37): C,
70.41; H, 3.08; N, 14.28. Found C, 70.56; H, 3.01; N, 14.14%.

Synthesis of 21a,b and 22.

A mixture of the heteroarylhydrazonalas 12 (0.96g, 2 mmol) and 2-cyanoacetamide 19a, or
3-oxo-3-phenylpropanenitrile 19b, or ethylcyanoactate 19c (2 mmol) in acetic acid (20 mL)
containing ammonium acetate (1 g) was refluxed for 4 h. then allowed to cool down to room
temperature and poured onto ice cold water. The formed precipitate was collected by
filtration washed with water and recrystallized from DMF to give pure 21a,b and 22.
2-(1,5-Dimethyl-3-oxo-2-phenyl-2,3-dihydro-1H-pyrazol-4-yl)-3-oxo-6-(3-oxo-3H-
benzo[f] chromene-2-carbonyl)-2,3-dihydropyridazine-4-carboxamide (21a): Orange
solid (78%); m.p. 247-9 °C (DMF); IR: v 1644,1669, 1690, 1708, 1726(5C=O), 3189,
3358(NH2) cm-1; 1H NMR: δ 2.34(3H, s, CH3), 3.39(3H, s, NCH3), 6.66-8.12(11H, m, Ar-H),
8.47(1H, s, pyridazinone-H4), 9.25(1H, s, Coumarin-H4), 11.34(2H, s, br, NH2); MS m/z
(%): 548(M+ + 1, 11), 547(M+, 100), 139(37), 80(67), 55(54). Anal. Calcd for C30H21N5O6
(547.52): C, 65.81; H, 3.87; N, 12.79. Found C, 65.59; H, 3.84; N, 12.66%.

4-Benzoyl-2-(1,5-dimethyl-3-oxo-2-phenyl-2,3-dihydro-1H-pyrazol-4-yl)-6-(3-oxo-3H-
benzo[f]chromene-2-carbonyl)pyridazin-3(2H)-one (21b): Yellow solid (87%); m.p. 288
°C (DMF); IR: v 1638,1657, 1697, 1712, 1735(5C=O) cm-1 ; 1H NMR: δ 2.36(3H, s, CH3),
3.42(3H, s, NCH3), 6.62-8.26(17H, m, Ar-H), 9.32(1H, s, Coumarin-H4); MS m/z (%):
608(M+, 7), 195(19), 144(100), 115(70), 63(31). Anal. Calcd for C36H24N4O4 (608.60): C,
71.05; H, 3.97; N, 9.21. Found C, 71.00; H, 3.92; N, 9.02%.

Ethyl 5-((1,5-dimethyl-3-oxo-2-phenyl-2,3-dihydro-1H-pyrazol-4-yl)diazenyl)-2-
hydroxy-6-(3-oxo-3H-benzo[f]chromen-2-yl)nicotinate (22):
Yellow solid (84%); m.p. 185-7 °C (DMF); IR: v 1657, 1709, 1727(3C=O), 3129-3414(OH)
cm-1 ; 1H NMR: δ 1.33(3H, t, J = 7.8 Hz, CH2CH3), 2.30(3H, s, CH3), 3.41(3H, s, NCH3),
4.29(2H, q, J = 7.8 Hz, CH2CH3), 6.78-8.14(13H, m, Ar-H, OH), 9.29(1H, s, Coumarin-
H4); MS m/z (%): 576(M+ + 1, 2), 575(M+, 2), 198(8), 144(100), 115(97), 55(11). Anal.
Calcd for C32H25N5O6 (575.57): C, 66.78; H, 4.38; N, 12.17. Found C, 66.54; H, 4.18; N,
12.12%.

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Reaction of 12 with -haloketones 23a,b

A mixture of 12 (0.96g, 2 mmol) and chloroacetone 23a or phenacyl bromide 23b (2 mmol)
in ethanol (30 mL) containing K2CO3 (0.5 g) was stirred at room temperature for 6h
(monitored by TLC). The reaction mixture was poured onto ice-water. The solid, so formed,
was collected by filtration and crystallized DMF to give 24a,b.

5-Acetyl-1',5'-dimethyl-3-(3-oxo-3H-benzo[f]chromene-2-carbonyl)-2'-phenyl-1'H-[1,4'-
bipyrazol]-3'(2'H)-one (24a): Yellow solid (73%); m.p. 246 °C (DMF); IR: v 1658, 1669,
1716, 1726(4C=O) cm-1 ; 1H NMR: δ 2.13(3H, s, CH3CO), 2.37(3H, s, CH3), 3.48(3H, s,
NCH3), 6.57-8.18(12H, m, Ar-H), 9.23(1H, s, Coumarin-H4); MS m/z (%): 518(M+, 33),
234(30), 195(79), 81(80), 55(100). Anal. Calcd for C30H22N4O5 (518.52): C, 69.49; H, 4.28;
N, 10.81. Found C, 69.25; H, 4.28; N, 10.59%.

5-Benzoyl-1',5'-dimethyl-3-(3-oxo-3H-benzo[f]chromene-2-carbonyl)-2'-phenyl-1'H-
[1,4'-bipyrazol]-3'(2'H)-one (24b): Yellow solid (77%); m.p. 279 °C (DMF); IR: v 1643,
1678, 1717, 1732(4C=O) cm-1; 1H NMR: δ 2.37(3H, s, CH3), 3.48(3H, s, NCH3), 6.68-
8.14(17H, m, Ar-H), 9.26(1H, s, Coumarin-H4); MS m/z (%): 580(M+, 33), 234(30), 195(79),
81(80), 55(100). Anal. Calcd for C35H24N4O5 (580.59): C, 72.40; H, 4.17; N, 9.65. Found C,
72.29; H, 4.11; N, 9.54%.

Pharmacology
Anticancer activity
The anticancer activity of the synthesized compounds was determined against the human
breast cancer cell line (MCF-7) and the liver carcinoma cell line (HEPG-2), using the 3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and doxorubicin was
used as a reference drug. Data generated were used to plot a dose–response curve of which
the concentration of test compounds required to kill 50% of cell population (IC50) was
determined. Cytotoxic activity was expressed as the mean IC50 of three independent
experiments (Table 1). The method applied is similar to that reported by Vijayan et al.28 using
Crystal violet stain (1%). Cells were seeded in 96-well plates at a cell concentration 1 x104
cells per well in 100 ml of growth medium. Fresh medium containing different concentrations
of the test sample was added after 24 h of seeding.

Serial twofold dilutions of the tested chemical compound were added to confluent cell
monolayers, flat-bottomed microtiter plates using a multichannel pipette. The microliter

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plates were incubated at 37 oC in a humidified incubator with 5% CO2 for 48 h. Three wells
were used for each concentration of the test sample. Control cells were incubated without test
sample with DMSO. After incubation of the cells for 24 h at 37 oC, various concentrations of
sample (50, 25, 12.5, 6.25, 3.125, and 1.56 g) were added, and the incubation was continued
for 48 h and the viable cell yield was determined by a colorimetric method. After the end of
the incubation period, media were aspirated and the crystal violet solution was added to each
well for at least 30 min. The stain was removed and the plates were rinsed using tap water
until all excess stain is removed. Acetic acid (30%) was then added to all wells and mixed
thoroughly, and then the absorbance of the plates was measured after gently shaken on
Microplate reader, using a test wavelength of 490 nm. All results were corrected for
background absorbance detected in wells without added stain. Treated samples were
compared with the cell control in the absence of the tested compounds. All experiments were
carried out in triplicate. The cell cytotoxic effect of each tested compound was calculated
(Table 1).

CONCLUSIONS
In conclusion, The results of the present study indicate that the sodium salt of 2-(3-
hydroxyacryloyl)-3H-benzo[f]chromen-3-one 2 is useful precursor for the synthesis of
different functionalized novel 2-(heteroaryl)-3H-benzo[f]chromen-3-ones.

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