Bis-(1H-2-benzopyran-1-one) derivatives: Synthesis and antimicrobial evaluation

Article abstract of DOI:10.1007/s12039-013-0529-0

The aim of the present study was to synthesize isocoumarin heterocycles and to elucidate the potential role of these compounds as biological active agents. A new series of isocoumarin derivatives containing two six-membered lactone rings is reported. 3-Aroyl-substituted isocoumarins (3) obtained by condensing 2- carboxy benzaldehyde (1) with bromoacetophenone derivatives (2) was further reacted with different aromatic aldehydes (4) affording bis-(1H-2-benzopyran-1- one) derivatives (5). This short review compiles examples of most promising antibacterial, antifungal and analgesic bis-(1H-2-benzopyran-1-one) derivatives. The products were characterized on the basis of analytical and spectral (IR, 1HNMR, C13NMR, Mass) data. The biological activity study revealed that all compounds showed promising activities and bis-(1H-2-benzopyran-1-one) derivatives (5) were found to be more active than 3-aroyl-substituted isocoumarins (3). Indian Academy of Sciences.

Full text of DOI:10.1007/s12039-013-0529-0

c
J. Chem. Sci. Vol. 125, No. 6, November 2013, pp. 1535–1542. ꢀ Indian Academy of Sciences.
Bis-(1H-2-benzopyran-1-one) derivatives: Synthesis
and antimicrobial evaluation
POONAM KUMAR KOPPULAa and NALINI PUROHITb,∗
aSchool of Science and Education, Navrachana University, Vadodara 391 410, India
bDepartment of Chemistry, Faculty of Science, The Maharaja Sayajirao University of Baroda,
Vadodara 390 002, India
e-mail: nalinipurohit22@gmail.com
MS received 20 May 2013; revised 4 September 2013; accepted 5 September 2013
Abstract. The aim of the present study was to synthesize isocoumarin heterocycles and to elucidate the poten-
tial role of these compounds as biological active agents. A new series of isocoumarin derivatives containing
two six-membered lactone rings is reported. 3-Aroyl-substituted isocoumarins (3) obtained by condensing 2-
carboxy benzaldehyde (1) with bromoacetophenone derivatives (2) was further reacted with different aromatic
aldehydes (4) affording bis-(1H-2-benzopyran-1-one) derivatives (5). This short review compiles examples of
most promising antibacterial, antifungal and analgesic bis-(1H-2-benzopyran-1-one) derivatives. The products
1
were characterized on the basis of analytical and spectral (IR, HNMR, C13NMR, Mass) data. The biologi-
cal activity study revealed that all compounds showed promising activities and bis-(1H-2-benzopyran-1-one)
derivatives (5) were found to be more active than 3-aroyl-substituted isocoumarins (3).
Keywords. 2-Carboxy benzaldehyde; 3-aroyl isocoumarin; bis-(1H-2-benzopyran-1-one) derivatives;
antibacterial activity; antifungal activity; analgesic activity.
1. Introduction
aminomethyl linkages at different positions in iso-
coumarin nucleus. Also, the structure activity rela-
tionships of isocoumarins have revealed that the mere
presence of isocoumarin moiety in any compound itself
is an essential feature of their pharmacological action.15
Furthermore, in recent decades, an increased inci-
dence of microbial infection has been observed as a
consequence of the growing number of immunocom-
promised patients and the frequent use of antibacterial
and cytotoxic drugs.
Despite the fact that the isocoumarin skeleton is
found in a variety of natural products, the dimeriza-
tion of isocoumarin itself has not been investigated.
There exist only few relevant examples in litera-
ture,16–18 related to the dimerization of coumarins and
its different derivatives along with their biological
applications.
Based on the wide spectrum of biological profile of
isocoumarins, and their increasing importance in phar-
maceutical and biological fields, and in continuation
of our ongoing research on biologically active hetero-
cycles, it was thought to be interest to accommodate
two identical isocoumarin moieties in a single molecu-
lar framework to synthesize some new heterocyclic
compounds with potential biological activity.
Among the 877 new molecule entities worldwide
introduced as drugs during 1981–2002, 61% can be
traced to or were inspired by natural products.1 These
include natural products, natural product derivatives,
synthetic compounds with natural product derived phar-
macophores and synthetic compounds designed on the
basis of knowledge gained from a natural product.2
Similarly, isocoumarins are an important class of
naturally occurring biologically active lactones, origi-
nating from a variety of natural sources, with multiple
biological activities.
Several articles dealing with the varied physiologi-
cal activities of isocoumarin derivatives have been
published, describing their anti-allergic, antimicrobial,
immunomodulatory, cytotoxic, antifungal, antiinflam-
matory and anti-angiogenic action. For the past many
years, the study of biological activities of isocoumarin
derivatives has been the aim of many researchers.3–11
In our earlier studies,12–14 we found an increase
in the bacteriological properties of isocoumarins on
introducing various substituted alkyl/acyl, aroyl and
∗For correspondence
1535

1536
Poonam Kumar Koppula and Nalini Purohit
7.42 (s, 1H, CH), 7.63–7.91 (m, 7H, aromatic protons),
2. Experimental
2.1 Materials and methods
8.44 (dd, 1H, CH); 13C NMR (CDCl3)δ 187 (C=O,
ketone), 164 (C=O, lactone), 142 (=C-O), 129 (C-Br),
132.3 (CH), 131.9 (CH), 136 (C), 115.6 (C), 136.5 (C),
128 (C), 130 (CH), 127.8 (CH), 133.6 (CH), 126 (CH);
ms: m/z: 328.97 (M+, 35%), 249(10), 174(19) and
146(100).
Reagents and solvents used in this study were of analyti-
cal grade and used without further purification. Melt-
ing points were determined in open capillaries and
are uncorrected. Purity of the compounds was checked
by Thin Layer Chromatography (TLC) on silica gel
GF254 plates using UV/iodine as visualizing agent
and Merk’s silica gel (60–120 mesh) was used for
column chromatographic purification. Infrared spec-
tra were recorded on Fourier Transform Infra Red
(FTIR) Perkin–Elmer spectrophotometer using potas-
2.2b 3-(4ꢁ-Hydroxy-benzoyl)-1H-2-benzopyran-1-one
(3b): White crystals, mp: 180◦C; 64.2% yield; IR
(KBr, v, cm−1): 1730 (γ lactone), 1610 (C=O), 3528
(OH); Anal. Calcd C16H10O4 (266.0 g): C, 72.18; H,
3.75; Found: C, 72.00; H, 3.18; 1H NMR (CDCl3)δ 5.57
(s, 1H, OH), 7.61 (s, 1H, CH), 7.04–7.86 (m, 7H, aro-
matic protons), 8.34 (d, 1H, CH); 13C NMR (CDCl3)δ
187 (C=O, ketone), 163 (C=O, lactone), 141.6
(=C-O), 163 (C-OH), 116 (CH), 131 (CH), 136 (C),
116 (C), 136.5 (C), 128 (C), 130 (CH), 127.8 (CH),
133.6 (CH), 126 (CH); ms: m/z: 266.04 (M+, 48%),
185(10), 145(60) and 121(100).
sium bromide optics. H NMR and 13C NMR spec-
1
tra were recorded on a Bruker spectrometer (400 MHz)
using Tetramethylsilane (TMS) as internal standard and
chemical shifts are given in ppm. Mass spectra were
obtained using Thermo Scientific Corporation, DSQ II
Mass Spectrometer. Elemental analyses were carried
out on Perkin–Elmer C, H, N, S Analyser (Model-
2400). Bromo acetophenone derivatives were prepared
by literature method.19 Antibacterial activity of newly
synthesized compounds was tested in vitro in bacte-
rial strains, Staphylococcus aureus (clinical strain) and
Escherichia coli (DH5 alpha) using serial agar dilution
(cup plate method)20 and Dimethylformamide (DMF)
were used as positive controls. Antifungal activity was
performed in vitro against fungal strains Fusarium pal-
lidoroseum and Colletotrichum capsici, using Potato
Dextrose Agar Medium (Poisoned Food Technique).21
Analgesic activity of the compounds was determined in
vivo by tail flick method.22 Gum acacia (2%) as control
and analgin drug as standard were used.
2.2c 3-(2ꢁ, 4ꢁ-Dihydroxy-benzoyl)-1H-2-benzopyran-
1-one (3c): Pinkish white crystals, mp: 116◦C; 44.6%
yield; IR (KBr, v, cm−1): 1733 (γ lactone), 1602
(C=O), 3556 (OH); Anal. Calcd C16H10O5 (282.0 g):
1
C, 68.08; H, 3.54; Found: C, 68.37; H, 4.08; H NMR
(CDCl3)δ 5.51 (s, 1H, OH), 5.87 (s, 1H, OH), 7.59
(s, 1H, CH), 6.74–7.72 (m, 6H, aromatic protons), 8.29
(d, 1H, CH); 13C NMR (CDCl3)δ 187 (C=O, ketone),
163 (C=O, lactone), 141 (=C-O), 165 (C-OH), 159
(C-OH), 108 (CH), 103 (CH), 132 (CH), 116.5 (C), 116
(C), 136.5 (C), 128.3 (C), 130 (CH), 127.6 (CH), 133.6
(CH), 126 (CH); ms: m/z: 282 (M+, 26%), 264(8),
238(15), 173(47), 146(65), 137(60) and 57(83).
2.2 General method for synthesis of 3-aroyl -1H-2-
benzopyran-1-one (3a–3f)
2.2d 3-(4ꢁ-Methoxy-benzoyl)-1H-2-benzopyran-1-one
(3d): Yellow crystals, mp: 134◦C; 62.8% yield; IR
(KBr, v, cm−1): 1728 (γ lactone), 1556 (C=O), 1083
(OCH3); Anal. Calcd C17H12O4 (280.0 g): C, 72.85; H,
4.28; Found: C, 73.24; H, 3.94; 1H NMR (CDCl3)δ 3.95
(s, 3H, OCH3), 7.42 (s, 1H, CH), 7.64–7.93 (m, 7H,
aromatic protons), 8.43 (dd, 1H, CH); 13C NMR
(CDCl3)δ 187 (C=O, ketone), 163 (C=O, lactone),
141.7 (=C-O), 168 (C-OCH3), 114.6 (CH), 131 (CH),
56 (CH3), 129 (C), 116 (C), 136.5 (C), 128 (C), 130
(CH), 127.8 (CH), 133.6 (CH), 126 (CH); ms: m/z:
280 (M+, 42), 252(54), 249(20), 145(74), 135(59) and
118(13).
2-Carboxy benzaldehyde (1 g, 0.006 mol) 1, 2-bromo-
1-(4-bromo-phenyl)-ethanone (1.85 g, 0.006 mol) 2,
K2CO3 (2.00 g, 0.012 mol) and ethyl methyl ketone
(10 ml) were taken in a round bottom flask and refluxed
for 10–12 h. Solvent was then removed, water added
and extracted with ethyl acetate. Solvent layer was first
washed with sodium bicarbonate and then with water
and dried over anh. Na2SO4. After removal of solvent,
the crude product was purified by column chromatogra-
phy using petroleum ether (60–80◦C)-ethyl acetate.
2.2a 3-(4ꢁ-Bromo-benzoyl)-1H-2-benzopyran-1-one
(3a): White crystals, mp: 171◦C; 76.0% yield; IR 2.2e 3-(Benzoyl)-1H-2-benzopyran-1-one (3e): White
(KBr, v, cm−1): 1735 (γ lactone), 1600 (C=O), 680 crystals, mp: 64◦C; 65.1% yield; IR (KBr, v, cm−1):
(C-Br); Anal. Calcd C16H9O3Br (328.9 g): C, 58.37; 1703 (γ lactone), 1629 (C=O); Anal. Calcd C22H14O3
H, 2.73; Found: C, 58.42; H, 3.06; 1H NMR (CDCl3)δ (326.0 g): C, 80.98; H, 4.29; Found: C, 81.29; H, 4.31;

Synthesis and biological evaluation of bis isocoumarins
1537
1H NMR (CDCl3)δ 7.42 (s, 1H, CH), 7.33–7.90 (m, (620.0 g): C, 75.48; H, 3.87; Found: C, 75.03; H, 4.17;
12H, aromatic protons), 8.41 (dd, 1H, CH); 13C NMR 1H NMR (CDCl3)δ 3.65 (s, 2H, OH), 5.85 (s, 1H,
(CDCl3)δ 187 (C=O, ketone), 163 (C=O, lactone), CH), 6.98–7.99 (m, 19H, aromatic protons), 8.27 (d,
141.6 (=C-O), 134 (CH), 129 (CH), 130 (CH), 137 2H, CH); 13C NMR (CDCl3)δ 163 (C=O, lactone),
(C), 116 (C), 136.7 (C), 128.1 (C), 130 (CH), 127.6 187 (C=O, ketone), 134.3 (=C-O), 163.1 (C-OH), 38.7
(CH), 133 (CH), 126.1 (CH); ms: m/z: 325 (M+−1, (CH), 116 (CH), 131 (CH), 129.3 (C), 125.5 (C), 136.4
45%), 300(62), 272(7), 221(21), 195(72), 174(27) and (C), 128.1 (C), 130 (CH), 127.3 (CH), 133.4 (CH),
146(95).
126.1 (CH), 125.4(C-H), 128 (CH), 129 (CH), 137.8
(C); ms: m/z: 620 (M+, 8%), 603(4), 577(26), 551(42),
509(8), 423(7), 368(38), 264(48) and 121(46).
2.2f 3-(Dibenzofuran-3-carbonyl)-1H-2-benzopyran-
1-one (3f): White crystals, mp: 110◦C; 65.6% yield;
IR (KBr, v, cm−1): 1720 (γ lactone), 1543 (C=O),
1253 (CO); Anal. Calcd C22H12O4 (340.0 g): C, 77.64;
H, 3.52; Found: C, 77.51; H, 4.01; 1H NMR (CDCl3)δ
7.76 (s, 1H, CH), 7.20–8.00 (m, 10H, aromatic pro-
tons), 8.45 (d, 1H, CH); 13C NMR (CDCl3)δ 187
(C=O, ketone), 163 (C=O, lactone), 141(=C-O),
132.8 (C), 113 (CH), 124.4 (CH), 122 (CH), 156 (C),
133.8 (C), 145 (C), 106.9 (C), 121 (CH), 123 (CH), 124
(CH), 111.8 (CH), 116 (C), 136.5 (C), 128 (C), 130
(CH), 127.8 (CH), 133.6 (CH), 126 (CH); ms: m/z:
340 (M+, 12%), 312(31), 174(3), 168(10) and 146(82).
2.3c 4,
4ꢁ-(4-Nitrobenzylidene)-bis-[3-(4ꢁ-hydroxy
benzoyl)-1H-2-benzopyran-1-one] (5c): White crys-
tals, mp: 198 ◦C; 68.2% yield; IR (KBr, v, cm−1):
1701(γ lactone), 1664 (C=O), 3428 (OH), 1486
(NO2); Anal. Calcd C39H23O10N (665.0 g): C, 70.37;
H, 3.45; N, 2.10; Found: C, 70.71; H, 3.92; N, 2.53; 1H
NMR (CDCl3)δ 6.70 (s, 1H, CH), 7.54–8.19 (m, 18H,
aromatic protons), 8.27 (d, 2H, CH) 9.35 (s, 2H, OH);
13C NMR (CDCl3)δ 163 (C=O, lactone), 187 (C=O,
ketone), 134.3 (=C-O), 163.1 (C-OH), 38.5 (CH),
116.2 (CH), 131.1 (CH), 129.1 (C), 125.7 (C), 136.5
(C), 128 (C), 130 (CH), 127.3 (CH), 133.6 (CH), 126.1
(CH), 145.4(C-NO2), 123 (CH), 130 (CH), 143.8 (C);
ms: m/z: 663 (M+−2, 2%), 619(8), 525(1), 479(2),
405(1), 266(30), 145(28) and 121(100).
2.3 General method for synthesis of
bis-(1H-2-benzopyran-1-one) derivatives (5a–5m)
A mixture of 3-aroyl isocoumarin 3b (1 g, 0.0037 mol)
was dissolved in ethanol and heated on water bath to
get a clear solution. p-chlorobenzaldehyde 4 (0.264 g,
0.0018 mol) was added to this hot solution and refluxed
for 18–20 h at 80◦C. After the reaction was over, the
solvent was distilled off and product recrystallized from
ethanol.
2.3d 4, 4ꢁ-(4-Methoxybenzylidene)-bis-[3-(4ꢁ-hydroxy
benzoyl)-1H-2-benzopyran-1-one] (5d): Yellow crys-
tals, mp: 200◦C; 72.0% yield; IR (KBr, v, cm−1): 1683
(γ lactone), 1624 (C=O), 3315 (OH), 1246 (OCH3);
Anal. Calcd C40H26O9 (650.0 g): C, 73.84; H, 4.00;
1
Found: C, 74.21; H, 4.38; H NMR (CDCl3)δ 3.84 (s,
3H, OCH3), 7.41 (s, 1H, CH), 7.36–8.01 (m, 18H, aro-
matic protons), 8.35 (d, 2H, CH) 8.77 (s, 2H, OH);
13C NMR (CDCl3)δ 163 (C=O, lactone), 187 (C=O,
ketone), 134.3 (=C-O), 163.1 (C-OH), 38.5 (CH),
116.2 (CH), 131.1 (CH), 129.1 (C), 125.7 (C), 136.5
(C), 128 (C), 130 (CH), 127.3 (CH), 133.6 (CH), 126.1
(CH), 159(O-CH3), 56 (C – OCH3), 114 (CH), 130
(CH), 130 (C); ms: m/z: 649 (M+−1, 2%), 616 (1), 588
(3), 560 (5), 529(2), 406(9), 383(13), 329(10), 280(42),
145(24) and 135(84).
2.3a 4, 4ꢁ- (4-Chlorobenzylidene)-bis-[3-(4ꢁ-hydroxy
benzoyl)-1H-2-benzopyran-1-one] (5a): Pale white
crystals, mp: 200◦C; 74.9% yield; IR (KBr, v, cm−1):
1710 (γ lactone), 1495 (C=O), 3112 (OH); Anal. Calcd
C39H23O8Cl (654.5 g): C, 71.61; H, 3.51; Found: C,
1
71.95; H, 3.14; H NMR (CDCl3)δ 5.75 (s, 1H, CH),
6.90–8.10 (m, 18H, aromatic protons), 8.39 (d, 2H,
CH), 9.95 (s, 2H, OH); 13C NMR (CDCl3)δ 163 (C=O,
lactone), 186 (C=O, ketone), 134 (=C-O), 163 (C-OH),
39 (CH), 116 (CH), 131 (CH), 129 (C), 126 (C), 137
(C), 128.3 (C), 130 (CH), 127.6 (CH), 133.6 (CH), 126
(CH), 130.6 (C-Cl), 129 (CH), 130 (CH), 135.8 (C);
ms: m/z: 480(7%), 410(5), 390(16), 375(24), 298(42),
266(50), 145(27), 121(49), 118(19) and 77(61).
2.3e 4, 4ꢁ-(4-Nitrobenzylidene)-bis-[3-(4ꢁ-bromo benzoyl)-
1H-2-benzopyran-1-one] (5e): Yellow crystals, mp:
144◦C; 68.2% yield; IR (KBr, v, cm−1): 1720 (γ lac-
tone), 1543 (C=O), 1446 (NO2), 526 (C-Br); Anal.
Calcd C39H21O8NBr2 (776.8 g): C, 59.18; H, 2.65; N,
1
2.3b 4, 4ꢁ-(Benzylidene)-bis-[3-(4ꢁ-hydroxy benzoyl)- 1.77; Found: C, 58.97; H, 2.74; N, 2.05; H NMR
1H-2-benzopyran-1-one] (5b): White crystals, mp: (CDCl3)δ 7.50 (s, 1H, CH), 7.31–8.10 (m, 18H, aro-
197◦C; 70.0% yield; IR (KBr, v, cm−1): 1718 (γ lac- matic protons), 8.44 (d, 2H, CH); 13C NMR (CDCl3)δ
tone), 1519 (C=O), 3252 (OH); Anal. Calcd C39H24O8 163 (C=O, lactone), 187 (C=O, ketone), 134.3

1538
Poonam Kumar Koppula and Nalini Purohit
(=C-O), 129 (C-Br), 38.6 (CH), 132.2 (CH), 131.8 130.1 (CH), 143.5 (C); ms: m/z: 693 (M+, 16%),
(CH), 135.7(C), 125.5 (C), 136.5 (C), 128.1 (C), 130 662(10), 525(10), 479(13), 405(15), 280(36), 145(29)
(CH), 127.6 (CH), 133.6 (CH), 126 (CH), 145 (C and 135(87).
–NO2), 123.3 (CH), 130 (CH), 144 (C); ms: m/z: 776.8
(M+,1%), 617(1), 571(2), 540(12), 447.9(17), 435(3),
407(2), 328.9(48), 187(20), 183.9(51), 146(45), and
77(75).
2.3i 4, 4ꢁ-(4-Hydroxybenzylidene)-bis-[3-(4ꢁ-methoxy
benzoyl)-1H-2-benzopyran-1-one] (5i): Yellow crys-
tals, mp: 90◦C; 61.7% yield; IR (KBr, v, cm−1):
1735 (γ lactone), 1600 (C=O), 3189 (OH), 1163
2.3f 4, 4ꢁ-(4-Hydroxybenzylidene)-bis-[3-(4ꢁ-bromo (OCH3); Anal. Calcd C41H28O9 (664.0 g): C, 74.09; H,
benzoyl-1H-2-benzopyran-1-one] (5f): Yellow crys- 4.21; Found: C, 73.86; H, 4.65; H NMR (CDCl3)δ
1
tals, mp: 150◦C; 69.0% yield; IR (KBr, v, cm−1): 1700 3.99 (s, 6H, OCH3), 7.35 (s, 1H, CH), 7.04–7.79 (m,
(γ lactone), 1643 (C=O), 3226 (OH), 580 (C-Br); Anal. 18H, aromatic protons), 8.39 (dd, 2H, CH) 12.70 (s,
Calcd C39H22O7Br2 (761.8 g): C, 61.43; H, 2.88; Found: 1H, OH); 13C NMR (CDCl3)δ 163 (C=O, lactone), 187
C, 61.72; H, 3.12; 1H NMR (CDCl3)δ 6.03 (s, 1H, OH), (C=O, ketone), 134.3 (=C-O), 167.8 (C-OCH3), 56
7.51 (s, 1H, CH), 7.30–8.15 (m, 18H, aromatic pro- (O–CH3), 38.7 (CH), 114.5 (CH), 130.7 (CH), 129 (C),
tons), 8.38 (d, 2H, CH); 13C NMR (CDCl3)δ 163 (C=O, 125.8 (C), 137(C), 128 (C), 130.2 (CH), 127.6 (CH),
lactone), 187 (C=O, ketone), 134.3 (=C-O), 128.9 134 (CH), 126.2 (CH), 154 (CH), 115 (CH), 130.5
(C-Br), 33 (CH), 132.3 (CH), 131.9 (CH), 135 (C), 126 (CH), 130.3 (C); ms: m/z: 662 (M+−2, 3%), 647(1),
(C), 137 (C), 128.3 (C), 130 (CH), 127.6 (CH), 133.4 585(19), 508(11), 497(14), 384(24), 392(30), 145(36)
(CH), 126 (CH), 154 (C–OH), 116 (CH), 130.5 (CH), and 135(49).
131 (C); ms: m/z: 761.8 (M+, 1%), 602(4), 585(6),
508(8), 432.9(15), 392(25), 183.9(10), 146(67) and
118(43).
2.3j 4, 4ꢁ-(4-Methoxybenzylidene)-bis-[3-(4ꢁ-methoxy
benzoyl)-1H-2-benzopyran-1-one] (5j): Yellow crystals,
mp: 134◦C; 60.0% yield; 1735 (γ lactone), 1626
2.3g 4, 4ꢁ-(Benzylidene)-bis-[3-(4ꢁ-bromo benzoyl)- (C=O), 1258 (OCH3); Anal. Calcd C42H30O9 (678.0 g):
1
1H-2-benzopyran-1-one] (5g): White crystals, mp: C, 74.33; H, 4.42; Found: C, 74.47; H, 4.68; H NMR
176◦C; 62.7% yield; IR (KBr, v, cm−1): 1689 (γ (CDCl3)δ 4.16 (s, 9H, OCH3), 7.45 (s, 1H, CH), 7.31–
lactone), 1652 (C=O), 632 (C-Br); Anal. Calcd 7.85 (m, 18H, aromatic protons), 8.40 (d, 2H, CH);
C39H22O6Br2 (745.8 g): C, 62.75; H, 2.94; Found: C, 13C NMR (CDCl3)δ 163 (C=O, lactone), 187 (C=O,
1
63.18; H, 3.36; H NMR (CDCl3)δ 7.41 (s, 1H, CH), ketone), 134.3 (=C-O), 167.4 (C-OCH3), 56.2 (O –
7.44–8.15 (m, 18H, aromatic protons), 8.45 (d, 2H, CH3), 39 (CH), 114.6 (CH), 130.7 (CH), 129 (C), 125.5
CH); 13C NMR (CDCl3)δ 163.2 (C=O, lactone), 186.8 (C), 136.5 (C), 128.3 (C), 130 (CH), 127.6 (CH), 133.6
(C=O, ketone), 134 (=C-O), 128.7 (C-Br), 32.6 (CH), (CH), 126 (CH), 158 (C–OCH3), 57 (O-CH3), 114
132 (CH), 131.9 (CH), 136 (C), 125.8 (C), 136.5 (C), (CH), 130.2 (CH), 130 (C); ms: m/z: 678 (M+, 17%),
128.3 (C), 130 (CH), 128 (CH), 133.4 (CH), 126.1 398(2), 280(27), 146(70), 135(41) and 118(52).
(CH), 125.5(CH), 128.3 (CH), 129 (CH), 137.7 (C);
ms: m/z: 745.8 (M+, 4%), 665.9(7), 509(1), 481(29),
376(11), 483.9(5), 416.9(32), 328.9(31), 183.9(41) and
146(60).
2.3k 4, 4ꢁ-(4-Nitrobenzylidene)-bis-[3-(2ꢁ, 4ꢁ-dihyroxy
benzoyl)-1H-2-benzopyran-1-one] (5k): Yellow crys-
tals, mp: 75◦C; 34.4% yield; 1702 (γ lactone),
1682 (C=O), 3320 (OH), 1590 (NO2); Anal. Calcd
2.3h 4,
4ꢁ-(4-Nitrobenzylidene)-bis-[3-(4ꢁ-methoxy C39H23O12N (697.0 g): C, 67.14; H, 3.29; N, 2.00;
benzoyl)-1H-2-benzopyran-1-one] (5h): Yellow crys- Found: C, 66.29; H, 3.57; N, 1.84; 1H NMR (CDCl3)δ
tals, mp: 128◦C; 54.3% yield; IR (KBr, v, cm−1): 7.42 (s, 1H, CH), 6.82–8.19 (m, 16H, aromatic pro-
1710 (γ lactone), 1682 (C=O), 1065 (OCH3), 1486 tons), 8.46 (d, 2H, CH), 11.60 (s, 2H, OH), 11.80
(NO2); Anal. Calcd C41H27O10N (693.0 g): C, 70.99; (s, 2H, OH) ; 13C NMR (CDCl3)δ 163 (C=O, lac-
H, 3.89; N, 2.02; Found: C, 71.23; H, 4.07; N, 1.94; tone), 187 (C=O, ketone), 134.3 (=C-O), 165 (C-
1H NMR (CDCl3)δ 3.74 (s, 6H, OCH3), 7.55 (s, 1H, OH), 160 (C-OH), 33 (CH), 108.8 (CH), 103.4 (CH),
CH), 7.00–7.90 (m, 18H, aromatic protons), 8.20 (d, 132 (CH), 116.5 (C), 125.5 (C), 136.5 (C), 128.3
2H, CH); 13C NMR (CDCl3)δ 163 (C=O, lactone), (C), 130 (CH), 127.6 (CH), 133.6 (CH), 126 (CH),
187 (C=O, ketone), 134.3 (=C-O), 168 (C-OCH3), 56 145.4 (C –NO2), 123.5 (CH), 130.1 (CH), 143.9 (C);
(O–CH3), 38.8 (CH), 115 (CH), 130.8 (CH), 129 (C), ms: m/z: 698 (M++1, 6%), 680(23), 617(8), 558(20),
125.5 (C), 136.5 (C), 128.3 (C), 130 (CH), 127.6 (CH), 540(34), 453(27), 415(20), 264(48), 173(39), 146(43)
133.6 (CH), 126 (CH), 145.5 (C –NO2), 123.3 (CH), and 137(44).

Synthesis and biological evaluation of bis isocoumarins
1539
2.3l 4, 4ꢁ-(4-Hydroxybenzylidene)-bis-[3-(2ꢁ, 4ꢁ- K2CO3 for 10–12 h, taking ethyl methyl ketone as sol-
dihydroxy benzoyl)-1H-2-benzopyran-1-one] (5l): White vent to get 3-aroyl isocoumarins (figure 1). Charac-
crystals, mp: 126◦C; 38.7% yield; 1742 (γ lactone), terization of synthesized compounds was done by IR
1706 (C=O), 3310 (OH); Anal. Calcd C39H25O11 and NMR spectral studies. All the compounds in 3a–
(669.0 g): C, 69.95; H, 3.73; Found: C, 70.21; H, 3.70; f show absorption at 1735–1700 cm−1 for lactonic car-
1H NMR (CDCl3)δ 7.47 (s, 1H, CH), 6.81–7.79 (m, bonyl and 1629–1543 cm−1 for ketonic carbonyl. Pres-
17H, aromatic protons), 8.38 (d, 2H, CH), 9.84 (s, 1H, ence of C-Br in 3a and –OH group in 3b and 3c is
OH), 12.53 (s, 2H, OH), 12.70 (s, 2H, OH) ; 13C NMR confirmed by signals at 680 cm−1 and 3528–3556 cm−1
(CDCl3)δ 163 (C=O, lactone), 187 (C=O, ketone), (br.), respectively. NMR spectrum of 3a–3d shows
134.3 (=C-O), 165 (C-OH), 160 (C-OH), 32 (CH), signals at δ 7.04–8.00 (m, 9H) for vinylic as well as
109 (CH), 103 (CH), 132.5 (CH), 117 (C), 125.5 (C), aromatic protons, at δ 3.95 (s, OCH3, 3H), with the
136.5 (C), 128.3 (C), 130 (CH), 127.6 (CH), 133.6 proton at 8th position showing a characteristic dou-
(CH), 126 (CH), 154 (C-OH), 115.5 (CH), 130.5 (CH), blet at around δ 8.34–8.44. Mass spectrum of 3a (MW
130.4 (C); ms: m/z: 668 (M+−1, 1%), 618 (2), 541(6), 328.9 g) shows M+ peak at m/z 328.97, base peak
436(16), 387(45), 331(22), 282(17), 173(40), 146(57) at m/z 146 for C9O2H6(isocoumarin moiety without
and 77(92).
any substitution) and m/z 174 for M+–C6H4Br. Mass
spectrum of 3d (MW 266) shows M+ peak at m/z
266.04, base peak at m/z 121 for M+-COC6H4OHand
m/z 146 for C9O2H6(isocoumarin moiety without any
substitution).
3-Aroyl isocoumarins 3 formed were then condensed
with different aryl aldehydes 4 in the ratio 2:1 in ethanol
to get bis-(1H-2-benzopyran-1-one) derivatives 5a–m
(figure 2). 5a shows IR signals for –OH, lactonic car-
bonyl and ketonic carbonyl, respectively. NMR spec-
trum of 5a shows signals at δ 5.75–7.50 (s, 1H, CH),
6.90–8.10 (m, 18H, aromatic protons), 8.25–8.45 (d,
2H, CH) 10.00–10.50 (s, 2H, OH). Mass spectrum of
5a gives m/z peak at 480, 410, 390, 375, 298, 266, 145,
121, 118 and 77.
2.3m 4, 4ꢁ-(4-Methoxybenzylidene)-bis-[3-(2ꢁ, 4ꢁ-
dihydroxy benzoyl)-1H-2-benzopyran-1-one] (5m):
Yellow crystals, mp: 144◦C; 35.2% yield; 1705 (γ lac-
tone), 1695 (C=O), 3568 (OH); Anal. Calcd C40H26O11
(683.0 g): C, 70.27; H, 3.95; Found: C, 69.84; H, 3.78;
1H NMR (CDCl3)δ 3.83 (s, 3H, OCH3), 5.79 (s, 4H,
OH), 7.42 (s, 1H, CH), 6.60–7.57 (m, 16H, aromatic
protons), 8.42 (d, 2H, CH); 13C NMR (CDCl3)δ 163
(C=O, lactone), 187 (C=O, ketone), 134.3 (=C-O),
164.5 (C-OH), 159.9 (C-OH), 33.3 (CH), 108.8 (CH),
103.4 (CH), 132 (CH), 116.5 (C), 125.5 (C), 136.5
(C), 128.3 (C), 130 (CH), 127.6 (CH), 133.6 (CH), 126
(CH), 159 (C–OCH3), 56 (O–CH3), 114 (CH), 130.2
(CH), 130 (C); ms: m/z: 683 (M+, 11%), 666 (8),
618(32), 541(10), 495(18), 439(5), 401(6) and 146(33).
3.2 Pharmacology
Some of the prepared compounds 3a–b, d, 5b–d
were tested in vitro against bacterial strains, Staphy-
lococcus aureus (Gram-positive) and Escherichia coli
(Gram-negative) using serial agar dilution (cup plate
method)20 and fungal strains of Fusarium pallidoro-
3. Results and discussion
3.1 Chemistry
2-Carboxy benzaldehyde was refluxed with different seum and Chaetonium using Potato Dextrose Agar
substituted bromo acetophenones in presence of anhy. medium (Poisoned Food Technique),21 respectively.
Figure 1. Synthetic pathway for the preparation of compounds 3a–f.

1540
Poonam Kumar Koppula and Nalini Purohit
Figure 2. Synthetic pathway for the preparation of compounds 5a–m.

Synthesis and biological evaluation of bis isocoumarins
1541
Table 1. Antimicrobial activity.
Zone of inhibition (mm)
% Growth of inhibition
Compound
S. aureus
E .coli
F. pallidoroseum Chaetonium
3a
3b
3d
5b
5c
5d
16
15
11
15
17
15
0
14
11
14
14
14
15
10
5
13.75
29.60
48.97
64.87
57.34
64.08
-
6.83
17.76
43.10
70.60
66.64
70.00
-
Control (DMF)
Ampicillin
15
-
-
Table 2. Analgesic activity.
Dose (mg/kg)
Average ( SE) reaction time (s)
time after drug treatment (min)
Compound
body weight
0
30
60
90
Control
Standard
3a
3b
3d
5b
5c
5d
50
50
50
50
50
50
50
50
3.01
3.09
1.32
2.46
3.62
4.00
3.53
3.41
3.20
5.25
3.28
3.00
3.97
4.53
4.20
4.73
3.10
7.75
3.69
3.04
4.01
5.98
5.68
5.05
3.02
9.00
4.77
3.88
5.35
7.44
6.52
6.93
The zone of inhibition of all compounds was found
Analgesic activity of the compounds was determined
to show moderate activity against Gram-negative i.e., by tail flick method22 on mice of either sex. Gum aca-
Escherichia coli bacteria and significant activity was cia (2%) was used as control, Analgin was the standard
seen with all compounds against Gram-positive bac- drug. 3-Aroyl isocoumarins fare badly in reaction time
teria S. aureus. Among all, excellent result was with when compared to the standard drug. The response time
5c where –NO2 group is substituted to isocoumarin is tremendous, almost equivalent to standard drug when
moiety apart from two –OH groups attached to aroyl two six-membered lactone rings are present together,
group. Promising effect was with 3a, where –Br was irrespective of the other groups present in them. This
substituted to aroyl group. Presence of hydroxyl group shows that the lactone ring in itself is useful in for
in aroyl moiety and –NO2 in isocoumarin moiety has biological applications.
enhanced antibacterial activity.
The results are shown in table 2.
The pattern of the result for antifungal activity of the
tested compounds was quite different from antibacterial
activity. Compound 3a–b,d were found to be weakly
or moderately active against both fungi where only one
isocoumarin moiety was present, but the compounds
5b–d where two isocoumarin moieties exist, showed
almost double the activity among all compounds. The
conclusion might be drawn that isocoumarin moiety
plays an important role in showing antifungal activity
rather than the substituent group present in it as seen in
antibacterial activity.
4. Conclusion
In conclusion, new bis-(1H-2-benzopyran-1-one)
derivatives were prepared from easily accessible
starting materials in a single step. The preliminary
in vitro test results of these compounds against the
four studied micro-organisms such as Staphylococcus
aureus, Escherichia coli, Fusarium pallidoroseum and
Chaetonium shows significant activity. All the tested
compounds showed good analgesic action also.
The results of antimicrobial screening of selected
new compounds are summarized in table 1.

1542
Poonam Kumar Koppula and Nalini Purohit
8. Ozcan S, Sahin E and Balci M 2007 Tetrahedron Lett.
Acknowledgements
48 2151
Authors are grateful to Sun Pharmaceutical Industries
Ltd. Baroda and SAIF, Punjab University for NMR
spectra analysis; and also to Prof. Anjana Desai,
Mrs. Aparna S of Microbiology Department, Prof. Arun
Arya, Head, Department of Botany, The Maharaja
Sayajirao University of Baroda for Antimicrobial
screening, and to Mr. G Paramesh, Gulbarga University
for analgesic activity.
9. Endringer D C, Guimaraes K G, Kondratyuk T P,
Pezzuto J M and Braga F C 2008 J. Nat. Prod. 71 1082
10. Bogdanov M G, Kandinska M I, Dimitrova D B,
Gocheva B T and Palamareva M D 2007 Z. Naturforsch
62 477
11. Kongsaeree P, Prabpai S, Sriubolmas N, Vongvein C and
Wiyakrutta S 2003 J. Nat. Prod. 66 709
12. Yadav P and Purohit N 2013 J. Chem. Sci. 125 165
13. Yadav P and Purohit N 2011 Indian J. Pharm. Sci. 73
171
14. Yadav P and Purohit N 2012 Der Pharmacia Lettre 4
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