Novel synthesis and cytotoxic activity of some chromeno[3,4-b]pyrrol-4(3H)- ones

Article abstract of DOI:10.1007/s10593-011-0701-8

1-Methylchromeno[3,4-b]pyrrol-4(3H)-one and 1-phenylchromeno[3,4-b]pyrrol- 4(3H)-one derivatives have been synthesized by cyclization of amides obtained by condensing α-halo ketones with 3-amino-chromenones, which show cytotoxic activity against lung, colon, and breast cancer cell lines.

Full text of DOI:10.1007/s10593-011-0701-8

Chemistry of Heterocyclic Compounds, Vol. 46, No. 12, March, 2011 (Russian Original Vol. 46, No. 12, December, 2010)
NOVEL SYNTHESIS AND CYTOTOXIC ACTIVITY
OF SOME CHROMENO[3,4-b]PYRROL-4(3H)-ONES
S. S. Soman1*, T. H. Thaker1, and R. A. Rajput2
1-Methylchromeno[3,4-b]pyrrol-4(3H)-one and 1-phenylchromeno[3,4-b]pyrrol-4(3H)-one derivatives
have been synthesized by cyclization of amides obtained by condensing α-halo ketones with 3-amino-
chromenones, which show cytotoxic activity against lung, colon, and breast cancer cell lines.
Keywords: benzopyrones, halo ketones, chromeno[3,4-b]pyrrol-4(3H)-ones, cytotoxic agents.
Benzo-α-pyrones (2H-chromen-2-ones), commonly known as coumarins are reported to possess a wide
range of biological activities [1–4]. Many natural and synthetic coumarins are reported to be cytotoxic agents [5].
Coumarins react with DNA by means of intercalation between two base pairs of DNA. There they form cross
linkage between two chains of DNA molecule, which is responsible for the carcinogenic and mutagenic effects.
3,4-Substituted coumarins have been reported to inhibit the proliferation of a number of human malignant cell
lines in vitro [6–7] and to possess anticoagulant and antithrombotic activities. These properties and our long and
continued interest in coumarin chemistry [8–10] encouraged us to synthesize new coumarin derivatives annelated
at positions 3 and 4 by the pyrrole ring. Literature survey reveals various methods for synthesis of such five-
membered heterocycle using metal-catalyzed reactions [11], which involves use of very costly palladium metal.
We report herein a very simple and novel method of synthesis of 3,4-annelated pyrrole derivatives of
benzopyrones. To the best of our knowledge, this method has not been used for the synthesis of the pyrrole
moiety.
3-Acetamidocoumarin (1) was condensed with different aldehydes in the presence of a base to get the
corresponding chalcones, but, instead, unreacted starting material was recovered. Therefore, acid hydrolysis of the
compound 1 was carried out to obtain 3-amino-2H-chromen-2-one (2a) (Scheme 1) [12]. It was then condensed
with cinnamoyl chloride in the presence of pyridine to produce the corresponding amide 3a.
Similarly, compound 2a was condensed with different derivatives of cinnamoyl chloride in pyridine to
give the corresponding 3-(3-arylacryloylamino)chromen-2-ones 3b–d. The structures of all these compounds
were confirmed by their IR and NMR spectra (Scheme 1).
_______
* To whom correspondence should be addressed, e-mail: shubhangiss@rediffmail.com, tirth1982@yahoo.co.in.
1Department of Chemistry, Faculty of Science, The Maharaja Sayajirao University of Baroda, Vadodara-390002,
India.
2National Institute of Pharmaceutical Education and Research, Thaltej, Ahmedabad, India.
__________________________________________________________________________________________
Translated from Khimiya Geterotsiklicheskikh Soedinenii, No. 12, pp. 1872-1878, December, 2010.
Original article received January 22, 2010, in revised form November 10, 2010.
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0009-3122/11/4612-1514©2011 Springer Science+Business Media, Inc.

Scheme 1
O
O
O
O
O
+
MeOH/H
ArCH
O
CHCCl
Pyridine
NHCOMe
NH2
1a
2a
O
N
O
UV,
Toluene
Ar
O
4
O
O
TFA,
DCM
NHCCH
O
CHAr
O
3a–d
NH
NH2NH2
Ar
O
5
No reaction
35 a Ar = Ph, b Ar = p-ClC6H4, c Ar = p-O2NC6H4, d Ar = p-MeOC6H4
Reaction of compounds 3a-d with hydrazine or photochemical cyclization [13] in a solvent like toluene to
get azetidinones 4 or heterocyclization in trifluoroacetic acid (TFA) [14] to get dihydropyridone 5 failed. Since no
desired product was obtained, the new methodology was used to synthesize pyrrole-annelated benzopyrones
(Scheme 2).
Scheme 2
O
O
R
NH2
2a, b
COCH2Br
abs. EtOH
O
O
R
ClCH2COMe
NHCH2COMe
6, 7
O
O
R
TFA
O
R
O
NHCH2C
O
R
O
O
8, 9
NH
TFA
NH
Me
10, 11
12, 13
2a,6,8,10,12 R = H; 2b,7,9,11,13 R = OH
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TABLE 1. Inhibition Compared to Control (Paclitaxel)
Inhibition, %
Cell line type
Colon
Compound
Concentration, ng/ml
Lung
Breast
250
500
31.8
54.3
66.5
85.0
54.3
72.2
78.1
91.1
33.5
52.2
61.5
80.2
49.8
68.8
79.5
88.2
38.1
53.5
75.7
87.2
56.1
77.7
83.4
93.4
10
1000
2000
250
11
500
1000
2000
Aminochromenones 2a,b were condensed with different α-halo ketones to get 3-(2-oxopropylamino)-
chromen-2-one (6), 7-hydroxy-3-(2-oxopropylamino)chromen-2-one (7), 3-(2-oxo-2-phenylethylamino)chromen-
2-one (8), and 7-hydroxy-3-(2-oxo-2-phenylethylamino)chromen-2-one (9). They were then cyclized in TFA to get
chromeno[3,4-b]pyrrol-4(3H)-ones 10–13 (Scheme 2). The structures of all compounds were confirmed by their
IR and NMR spectra. The disappearance of the signal at δ 4.9 for CH2 proton from the NMR spectra of 10-13
clearly indicated that cyclization took place. In conclusion, we found that the new methodology worked well for the
synthesis of new pyrrole derivatives of coumarin in good yields.
Cytotoxicity screening models provide important preliminary data to help select plant extracts with
potential anticancer properties for future work.
The cytotoxic activity study of compounds 10 and 11 was done using the MTT [3-(4,5-dimethylthiazol-
2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay against various cell lines of colon, lung, and breast cancer. It
was observed that both compounds show good activity against all the cell lines.
To conclude, this paper describes a simple and easy method to synthesize pyrrole derivatives of
benzopyrone, which show cytotoxic activity against lung, colon, and breast cancer cell lines.
EXPERIMENTAL
The melting points were taken in scientific open capillaries and are uncorrected. The IR spectra in KBr
1
were recorded on a Shimadzu IR-408 spectrophotometer. The H NMR spectra were recorded in CDCl3 or
DMSO-d6 on Bruker DRX-400 or Bruker DRX-300 FT NMR instruments using tetramethylsilane as an internal
standard. Mass spectra were recorded on a QP 5050A GC-MS spectrometer. Elemental analyses of compounds
were done on a Carlo Erba-1108 elemental analyzer. Acme Silica gel (mesh size 60–120) was used for column
chromatography.
3-Amino-2H-chromen-2-one (2a) was synthesized from compound 1a following a published procedure
[12].
3-Amino-7-hydroxy-2H-chromen-2-one (2b). A mixture of resorcaldehyde (5.0 g, 0.036 mol),
N-acetylglycine (3.6 g, 0.036 mol), acetic anhydride (15 ml, 0.14 mol), and sodium acetate (7.4 g, 0.9 mol) was
heated on a steam bath for 6 h to give light-yellow 7-hydroxy-N-(2-oxo-2H-chromen-3-yl)acetamide (1b), which
was recrystallised from alcoholwater, 1:1; mp 243°C. Refluxing of compound 1b with methanolic HCl (70 ml,
10%) gave 3-aminocoumarin 2b, which was recrystallized from alcohol and obtained as a light-yellow solid, mp
227°C, yield 52.2%.
Preparation of Amides 3a-d (General Method). To a mixture of compound 2a (1.61 g, 10 mmol),
triethylamine (1.4 ml, 10 mmol), and a catalytic amount of pyridine in dichloromethane (25 ml), different
1516

cinnamoyl chlorides (10 mmol) were added dropwise at 0–5°C, with constant stirring. The reaction mixture was
stirred at 0-5°C for 6 h. The organic phase was then partitioned at first with sat. NaHCO3 and then with water.
The organic portions were combined and dried over anhydrous Na2SO4 and filtered, and the solvent was
evaporated to give a crude product. It was recrystallized using ethanol–DMF, 8:2, to give different pure
cinnamides.
N-(2-Oxo-2H-chromen-3-yl)-3-phenylacrylamide (3a) (1.40 g, 48%) was obtained as a brown solid;
mp 226°C. IR spectrum, , cm–1: 1205, 1536, 1630, 1674, 1715, 3313. 1H NMR spectrum (300 MHz, DMSO-d6),
δ, ppm (J, Hz): 7.33-7.49 (6H, m, H-α,β, H-5–8); 7.62–7.74 (5H, m, H-2'–6'); 8.79 (1H, s, H-4); 9.94 (1H, s, NH).
13C NMR spectrum (100 MHz, DMSO-d6), , ppm: 115.5, 119.4, 121.5, 123.4, 124.6, 127.4, 127.6, 128.5, 129.1,
129.5, 134.5, 141.1, 149.4, 157.4, 165.0. Found, %: C 74.11; H 4.51; N 4.92. C18H13NO3. Calculated, %: C 74.23;
H 4.46; N 4.81.
3-(4-Chlorophenyl)-N-(2-oxo-2H-chromen-3-yl)acrylamide (3b) (1.69 g, 52%) was obtained as light-
brown solid; mp 257°C. Found, %: C 66.23; H 3.51; N 4.52. C18H12ClNO3. Calculated, %: C 66.36; H 3.69;
N 4.30.
3-(4-Nitrophenyl)-N-(2-oxo-2H-chromen-3-yl)acrylamide (3c) (1.88 g, 56%) was obtained as a yellow
solid; mp 261°C. IR spectrum, , cm–1: 1182, 1519, 1537, 1621, 1677, 1708, 3328. 1H NMR spectrum (400 MHz,
DMSO-d6), , ppm (J, Hz): 7.31–7.35 (2H, m, H-6,8); 7.42–7.46 (1H, d, J = 15.6, H-α); 7.48 (1H, m, H-7);
7.56-7.58 (1H, m, H-5); 7.69-7.73 (1H, d, J = 15.7, H-β); 7.77–7.79 (2H, d, J = 8.7, H-2',6'); 8.24–8.27 (2H, d,
J = 8.7, H-3',5'); 8.87 (1H, s, H-4); 9.85 (1H, s, NH). Found, %: C 64.16; H 3.52; N 8.40. C18H12N2O5. Calculated,
%: C 64.28; H 3.57; N 8.33.
3-(4-Methoxyphenyl)-N-(2-oxo-2H-chromen-3-yl)acrylamide (3d) (2.02 g, 63%) was obtained as a light-
brown solid; mp 271°C. Found, %: C 71.21; H 4.55; N 4.42. C19H15N4O. Calculated, %: C 71.10; H 4.67; N 4.36.
3-(2-Oxopropylamino)chromen-2-one (6). A mixture of compound 2a (1.61 g, 10 mmol), α-chloro-
acetone (0.80 ml, 10 mmol), and a catalytic amount of potassium iodide was refluxed for 5 h and then allowed to
reach room temperature. The precipitated crude product was filtered off, dried, and purified by column
chromatography using ethyl acetatepetroleum ether, 1:9, as eluent. The title compound 6 (0.89 g, 41%) was
obtained as a yellow solid; mp 210°C. IR spectrum, , cm–1: 745, 1216, 1357, 1458, 1534, 1629, 1702, 1720,
1
3329. H NMR spectrum (400 MHz, CDCl3), δ, ppm (J, Hz): 1.55 (3H, s, CH3); 4.24 (2H, s, CH2); 6.70 (1H, s,
H-8); 7.18-7.22 (1H, m, H-6); 7.25-7.30 (2H, m, H-5,7); 8.07 (1H, s, H-4). Found, %: C 66.42; H 5.01; N 6.51.
C12H11NO3. Calculated, %: C 66.36; H 5.06; N 6.45.
7-Hydroxy-3-(2-oxopropylamino)chromen-2-one (7). Compound 2b was used instead of compound 2a,
and the procedure was the same as that for compound 6. The title compound 7 (1.12 g, 48%) was obtained as an
orange solid; mp 232ºC. IR spectrum, , cm–1: 1127, 1153, 1165, 1243, 1289, 1460, 1508, 3224, 3349, 3630-2849
1
(broad). H NMR spectrum (400 MHz, CDCl3), δ, ppm (J, Hz): 2.64 (3H, s, CH3); 4.23 (2H, s, CH2); 6.69-6.79
(2H, m, H-6,8); 7.11-7.13 (1H, d, J = 8.4, H-5); 7.38 (1H, s, H-4); 9.33 (1H, s, NH); 11.82 (1H, s, OH). Found, %:
C 61.58; H 4.81; N 6.05. C12H11NO4. Calculated, %: C 61.80; H 4.72; N 6.00.
3-(2-Oxo-2-phenylethylamino)chromen-2-one (8). A mixture of compound 2a (1.61 g, 10 mmol) and
phenacyl bromide (1.98 g, 10 mmol) in absolute ethanol was refluxed for 5 h and then allowed to reach room
temperature. The precipitated crude product was filtered off, dried, and recrystallized from absolute ethanol. The
title compound 8 (1.56 g, 56%) was obtained as a yellow solid; mp 218°C. IR spectrum, , cm–1: 1508, 1606,
1
1628, 1705, 3328, 3403. H NMR spectrum (300 MHz, CDCl3), δ, ppm (J, Hz): 4.63–4.65 (2H, d, J = 4.8, CH2);
6.36-6.39 (1H, t, J = 4.8, NH); 7.25-7.29 (1H, dd, J = 2.7, J = 8.4, H-8); 7.32–7.39 (3H, m, H-5–7); 7.40–7.55 (5H,
m, Ar); 8.07 (1H, s, H-4). Found, %: C 73.06; H 4.69; N 5.28. C17H13NO3. Calculated, %: C 73.11; H 4.66; N 5.01.
7-Hydroxy-3-(2-oxo-2-phenylethylamino)chromen-2-one (9). Compound 2b was used instead of
compound 2a, and the procedure was the same as that for compound 8. The title compound 9 (1.80 g, 61%) was
obtained as a orange solid; mp 248°C. IR spectrum, , cm–1: 1499, 1615, 1684, 1728, 3405, 3652–2880 (broad).
1H NMR (400 MHz, CDCl3), δ, ppm (J, Hz): 4.63 (2H, d, J = 4.8, CH2); 5.72 (1H, t, J = 4.8, NH); 6.43 (1H, s, H-4);
1517

6.76-6.80 (2H, m, H-6,8); 7.17-7.19 (1H, d, J = 8.3, H-5); 7.52–7.70 (3H, m, H-3'–5'); 8.04-8.06 (2H, m, H-2',6');
9.39 (1H, s, OH). Found, %: C 69.03; H 4.44; N 4.82. C17H13NO4. Calculated, %: C 69.15; H 4.40; N 4.74.
Preparation of Compounds 10–13 (General Method). Trifluoroacetic acid was added in catalytic amount to
a stirred solution of compounds 6-9 (10 mmol) in acetic acid. The mixture was refluxed for 12 h and poured onto
crushed ice, and the crude product was filtered off, dried, and purified by gradient column chromatography using
ethyl acetate and petroleum ether as eluent. The pure product was eluted with ethyl acetatepetroleum ether, 1:9.
1-Methylchromeno[3,4-b]pyrrol-4(3H)-one (10) (1.35 g, 68%) was obtained as white crystals; mp 252°C.
1
IR spectrum, , cm–1: 707, 765, 1247, 1292, 1359, 1451, 1527, 1603, 1682, 1711, 3078, 3337, 3461. H NMR
spectrum (400 MHz, DMSO-d6), δ, ppm (J, Hz): 2.22 (3H, s, CH3); 7.27-7.75 (4H, m, H-69); 8.64 (1H, s, H-2);
9.49 (1H, s, NH). 13C NMR spectrum (100 MHz, DMSO-d6), , ppm: 23.8, 115.4, 119.3, 123.0, 124.2, 124.4,
127.1, 127.8, 128.8, 149.3, 157.7, 169.9. GC-mass spectrum (CI), m/z: 204 [M+4]+. Found, %: C 72.28; H 4.60;
N 6.95. C12H9NO2. Calculated, %: C 72.36; H 4.52; N 7.03.
7-Hydroxy-1-methylchromeno[3,4-b]pyrrol-4(3H)-one (11) (0.67 g, 31%) was obtained as a brown solid;
1
mp 258°C. IR spectrum, , cm–1: 1378, 1538, 1601, 1704, 3322, 3395, 2851–3751 (broad). H NMR spectrum
(400 MHz, CDCl3), , ppm (J, Hz): 2.20 (3H, s, CH3); 6.73-6.81 (2H, m, H-6,8); 7.31–7.33 (1H, d, J = 8.4, H-9);
13
8.55 (1H, s, H-2); 9.10 (1H, s, OH); 9.95 (1H, s, NH). C NMR spectrum (100 MHz, DMSO-d6), δ, ppm: 23.7,
101.9, 111.2, 113.3, 120.6, 124.9, 128.0, 151.0, 158.1, 159.1, 169.4. GC-mass spectrum (CI), m/z: 220 [M+4]+.
Found, %: C 66.61; H 4.12; N 6.44. C12H9NO3. Calculated, %: C 66.97; H 4.18; N 6.51.
1-Phenylchromeno[3,4-b]pyrrol-4(3H)-one (12) (1.33 g, 51%) was obtained as a light-brown solid; mp
178°C. Found, %: C 78.29; H 4.12; N 5.11. C17H11NO2. Calculated, %: C 78.16; H 4.21; N 5.36.
7-Hydroxy-1-phenylchromeno[3,4-b]pyrrol-4(3H)-one (13). Compound 13 (0.78 g, 28%) was obtained
1
as a red solid; mp 243°C. IR spectrum, , cm–1: 1505, 1607, 1733, 3356, 2872–3607 (broad). H NMR spectrum
(400 MHz, DMSO-d6), δ, ppm (J, Hz): 6.78 (1H, s, H-6); 7.41–7.58 (5H, m, H-2,8,3',4',5'); 7.91–7.93 (1H, d,
J = 8.5, H-9); 8.05–8.07 (2H, d, J = 8.7, H-2',6'); 9.42 (1H, s, NH); 10.82 (1H, s, OH). Found, %: C 73.69;
H 4.02; N 5.01. C17H11NO3. Calculated, %: C 73.64; H 3.97; N 5.05.
Biological Activity Study. The cell lines were procured from National Centre for Cell Science, Pune,
India. The cell lines HCT-15 for colon cancer (DMEM medium), A 549 for lung carcinoma (RPMI medium), and
MCF-7 for breast cancer (CMEM medium) were used for cytotoxic activity study.
MTT Assay [3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium Bromide]. Exponentially growing
cancer cells (HCT-15, A549 and MCF-7 cells) were harvested from 75 mm2 flask, and a stock cell suspension was
prepared. Then 5  104 cells/ml were seeded in a 96-well flat bottom tissue culture plate with 200 µl of complete
medium and incubated for 24 h. The compounds were dissolved in the minimum amount of DMSO and were
prepared in complete media without phenol red. All samples were first sterilized using Durapore polyvinylidene
difluoride membrane filters of 0.22 µm (Millipore, Ireland). Paclitaxel (5 µM) was used as positive control. Cells
were treated with different concentrations of compounds in 100 µl volume prepared in media without phenol red
and with positive control (Paclitaxel), and were incubated for 48 h. The cells in the control group received no
drug treatment. Each treatment was performed in three-well replicates. After the treatment, drug- containing
media was removed and washed twice with 100 µl of phosphate buffered saline. To each well of the 96-well
plate, 20 µl of MTT reagent (Merck, India) (stock: 5 mg/ml) was added and the wells incubated for 4 h at 37ºC.
Plates were shaken for 10 min and inverted with gentle tapping on tissue paper to remove the media. To solubilize
formazan crystals in the wells, 100 µl of 100% DMSO was added to each well. Plates were placed on a rotary
shaker (R 100/TW LUCKHAM, England) and agitated for 15-20 min.
The optical density (O.D.) was measured by an Enzyme Linked Immunosorbent assay Bio-Rad plate
reader at 570 nm with a reference wavelength of 630 nm. The O.D. of each well was read and expressed as
percentage cell viability (absorbance of treated wells / absorbance of control wells × 100). Results were expressed
as mean ± S.D. % cell viability values plotted against the crude extract concentrations. From the results, less than
50% survival at exposure time of 48 h was considered to be significant.
1518

The authors are thankful to the Head of Department of Chemistry for providing necessary facilities. The
authors are thankful to UGC, New Delhi, for funding. One of the authors (THT) is thankful to UGC, New Delhi,
for providing a fellowship.
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