untitled


European	Journal	of	Chemistry	3	(1)	(2012)	51‐56	

European	Journal	of	Chemistry	
ISSN	2153‐2249	(Print)	/	ISSN	2153‐2257	(Online)		2012	EURJCHEM	

DOI:10.5155/eurjchem.3.1.51‐56.500	

	
	

	

	

European	Journal	of	Chemistry	
Journal	homepage:	www.eurjchem.com	

	 	 	

Cyclocondensation,	antimicrobial	activity	and	semi‐empirical	AM1‐MO	
calculations	of	benzopyrone	derivatives	

Hafez	Mohamed	El‐Shaaer	

Department	of	Chemistry,	Faculty	of	Education,	Ain	Shams	University,	Roxy,	11711,	Cairo,	Egypt	

*Corresponding	author	at:	Department	of	Chemistry,	Faculty	of	Education,	Ain	Shams	University,	Roxy,	11711,	Cairo,	Egypt.		
Tel.:	+201.12627069;	fax:	+202.2581243.	E‐mail	address:	elshaaer@hotmail.com	(H.M.	El‐Shaaer).	

	
	

	 	

	 	 	
ARTICLE	INFORMATION	 	 ABSTRACT
Received:	13	August	2011	
Received	in	revised	form:	06	September	2011	
Accepted:	22	September	2011	
Online:	31	March	2012	

KEYWORDS	

	 An	 efficient	 synthesis	 of	 7‐amino‐9‐hydroxy‐6‐oxo‐6H‐benzo[c]chromene‐8‐carbonitrile
derivatives	 (2,6)	 and	 4‐methyl‐2‐oxo‐2H‐chromene‐3‐carbonitrile	 (5)	 via	 Claisen
condensation	 of	 2‐hydroxyacetophenones	 (1,3)	 with	 ethyl	 cyanoacetate	 in	 the	 presence	 of
sodium	metal	is	reported.	Reaction	of	5	with	thiosemicarbazide	gave	1‐(3‐cyano‐4‐methyl‐2‐
oxoquinolin‐1(2H)‐yl)thiourea	 (7).	 Treatment	 of	5	 with	 6,8‐dichloro‐4‐oxo‐4H‐chromene‐3‐
carboxaldehyde	 gave	 4‐(2‐(6,8‐dichloro‐4‐oxo‐4H‐chromen‐3‐yl)‐2‐hydroxyethyl)‐2‐oxo‐2H‐
chromene‐3‐carbonitrile	 (9).	 Further	 treatment	 of	 5	 with	 ethyl	 acetate	 followed	 by
condensation	with	6,8‐dichloro‐3‐formylchromone	gave	7‐((6,8‐dichloro‐4‐oxo‐4H‐chromen‐
3‐yl)methyleneamino)‐9‐hydroxy‐6H‐benzo[c]chromen‐6‐one	 (12).	 Structures	 of	 the
products	were	established	on	the	basis	of	elemental	analysis,	IR,	1H	and	13C	NMR,	mass	spectra
and	 semi‐empirical	 AM1‐MO	 calculations.	 The	 antimicrobial	 activities	 of	 the	 synthesized
products	were	also	studied.	

Synthesis	
Benzopyrone	
4‐Quinolinone	
Cyclocondensation	
AM1‐MO	calculation	
Antimicrobial	activity	

	
1.	Introduction	
	

Fused	 coumarins	 (2H‐1‐benzopyran‐2‐ones)	 comprise	 a	
very	 interesting	 class	 of	 compounds	 due	 to	 their	 significant	
antibacterial	 [1]	 and	 pharmacological	 activities	 [2].	 Also,	
chromone	 (4H‐1‐benzopyran‐4‐one)	 derivatives	 exhibit	
significant	 biological	 activities,	 such	 as	 antifungal	 [3,4],	
antimycobacterial	[5,6],	antialergic	[7],	antitumour	[8‐10],	and	
antiviral	[11‐12].	So,	and	with	the	expectative	to	find	biological	
activity,	 I	 decided	 to	 investigate	 the	 synthesis	 of	 some	 novel	
systems	 of	 coumarin	 derivatives	 bearing	 chromone	 moiety.	
Recently,	 the	 synthesis	 [13],	 photochemical	 [14]	 and	
theoretical	 [15]	 properties	 of	 chromone	 derivatives	 were	
largely	investigated.	

The	 aim	of	 the	present	paper	 is	 to	 investigate	 an	 efficient	
synthesis	of	coumarin	derivatives	containing	active	methyl	and	
cyano	groups	and	study	their	cyclocondensation	reactions	with	
ethyl	 cyanoacetate,	 thiosemicarbazide,	 ethyl	 acetate	 and	 6,8‐
dichloro‐4‐oxo‐4H‐chromene‐3‐carboxaldehyde.	 The	 anti‐
microbial	 activities	 for	 the	 prepared	 compounds	 were	
investigated.	Also,	semi‐empirical	AM1	and	Ab	Initio	(STO‐3G)	
molecular	 orbital	 calculations	 for	 the	 new	 compounds	 were	
studied	and	compared	with	their	experimental	data.	
	
2.	Experimental	
	
2.1.	Instrumentation	
	

The	 uncorrected	 melting	 points	 were	 determined	 in	 an	
open	capillary	tube	on	a	digital	Stuart	SMP‐3	apparatus.	1H/13C	
NMR	spectra	were	obtained	on	a	500/125	MHz	Jeol	Eca	or	on	a	
300/75.46	MHz	Varian	Mercury	VX‐300	NMR	spectrometer	 in	
DMSO‐d6	 with	 tetramethylsilane	 as	 an	 internal	 standard.	
Elemental	 analyses	 were	 performed	 on	 Vario	 El	 Elementar	
apparatus.	 IR	 spectra	 were	 recorded	 on	 FTIR	 Nicolet	 IS10	

spectrophotometer	(cm‐1),	using	KBr	disks.	Mass	spectra	were	
recorded	 on	 a	 Gas	 Chromatographic	 GCMSqp	 1000	 ex	
Shimadzu	 instrument	 at	 70	 eV.	 Thermodynamic	 data	 were	
obtained	 from	molecular	mechanical	 calculations	 on	 the	basis	
of	 the	 semi‐empirical	 AM1	 and	 Ab	 Initio	 (STO‐3G)	 methods	
with	the	HyperChem	8.03	computer	program.	
	
2.2.	Synthesis	
	

6,8‐Dichloro‐4‐oxo‐4H‐chromene‐3‐carboxaldehyde	 (8)	
was	 prepared	 according	 to	 [15].	 The	 other	 chemicals	 were	
purchased	from	the	suppliers	as	the	highest	purity	grade.	
	
2.2.1.	7‐Amino‐2,4‐dichloro‐9‐hydroxy‐6‐oxo‐6H‐benzo[c]	
chromene‐8‐carbonitrile	(2)	
	

A	solution	of	3,5‐dichloro‐2‐hydroxyacetophenone	(1)	(4.0	
g,	2	mmol)	in	ethyl	cyanoacetate	(22	cm3),	sodium	metal	(2	g,	8	
mmol)	 was	 added	 by	 small	 portion	 on	 the	 solution	 and	 the	
reaction	mixture	 was	 heated	 on	water‐bath	 for	 2	 h	 and	 then	
heated	 under	 reflux	 for	 1	 h.	 The	 product	 was	 treated	 with	
water	 and	 acidified	 with	 acetic	 acid.	 The	 solid	 obtained	 was	
filtered,	 and	 crystallized	 from	 acetic	 acid	 to	 give	 2	 as	 yellow	
crystals	(Scheme	1).	

7‐Amino‐2,4‐dichloro‐9‐hydroxy‐6‐oxo‐6H‐benzo[c]chrome	
ne‐8‐carbonitrile	(2):	Yield:	29%.	M.p.:	270‐272	°C.	FT‐IR	(KBr,	
cm‐1):	3350,	3260,	3197,	3064,	2981,	2209,	1736,	1682,	1616.	
1H	NMR	(300	MHz,	DMSO‐d6):	2.50	 (m,	3H,	CH,	CH2),	4.19	 (m,	
2H,	NH2,	exchangeable	with	D2O),	8.02‐8.09	(m,	2H,	H‐1	and	H‐
3),	 8.88	 (s,	 1H,	 exchangeable	 with	 D2O),	 9.19	 (s,	 1H,	
exchangeable	with	D2O).	13C	NMR	(75.46	MHz,	DMSO‐d6):	60.0,	
72.0,	 109.2,	 117.5,	 120.7,	 120.9,	 121.4,	 122.3,	 125.1,	 129.1,	
132.5,	 146.6,	 150.4,	 155.8,	 163.2,	 166.1.	 MS	 (EI,	 m/z	 (%)):	
322.4	(M+1,	15.9),	322.8	(M+2,	18.2).		

	



52	 El‐Shaaer	/	European	Journal	of	Chemistry	3	(1)	(2012)	51‐56	
	

	

Scheme	1	
	
	

	
	

Scheme	2	
	

	
Anal.	 calcd.	 for	 C14H6Cl2N2O3:	 C,	 52.36;	 H,	 1.88;	 N,	 8.72.	

Found:	C,	52.54;	H,	2.00,	N,	8.72%.	
	
2.2.2.	4‐Methyl‐2‐oxo‐2H‐chromene‐3‐carbonitrile	(5)	and	7‐
amino‐9‐hydroxy‐6‐oxo‐6H‐benzo[c]chromene‐8‐
carbonitrile	(6)	
	

Method	 A:	 Sodium	 metal	 (10	 g,	 43	 mmol)	 was	 added	 in	
small	portion	to	a	solution	of	2‐hydroxyacetophenone	(3)	(20.0	
g,	 15	 mmol)	 in	 ethyl	 cyanoacetate	 (110	 cm3).	 The	 reaction	
mixture	was	heated	on	water‐bath	 for	3	h,	and	then	cooled	 to	
room	temperature,	treated	with	ethanol	(30	cm3)	and	refluxed	
for	 1	h.	 The	 solid	 obtained	was	 filtered,	 and	 crystallized	 from	
cyclohexane	to	give	5	as	yellow	crystals,	Yield:	20%.	M.p.:	192‐
194	 °C	 (Scheme	 2	 and	 3).	 The	 ethanolic	 filtrate	 was	
concentrated	to	its	half	amount,	acidified	with	96%	acetic	acid	
and	diluted	with	water	 to	give	6	as	pall‐yellow	crystals,	Yield:	
30%.	M.p.:	237‐238	°C	(acetic	acid).		

4‐Methyl‐2‐oxo‐2H‐chromene‐3‐carbonitrile	(5):	FT‐IR	(KBr,	
cm‐1):	3060,	2920,	2228,	1723,	1600.	1H	NMR	(500	MHz,	DMSO‐
d6):	2.69	(s,	3H,	CH3),	7.44‐7.47	(m,	2H,	H‐6	and	H‐8),	7.76	(t,	J	=	
7.65	Hz,	1H,	H‐7),	7.95	(d,	J	=	8.45	Hz,	1H,	H‐5).	13C	NMR	(125	
MHz,	DMSO‐d6):	18.7	(‐),	101.8	(quat),	114.8	(quat),	117.5	(+),	
118.6	(quat),	125.8	(+),	127.6	(+),	135.8	(+),	153.2	(quat),	157.3	
(quat),	164.2	(quat).	MS	(EI,	m/z	(%)):	184	(M‐1,	13.7),	185	(M,	
100.0),	186	(M+1,	12.1).	Anal.	calcd.	 for	C11H7NO2:	C,	71.35;	H,	
3.81;	N,	7.56.	Found:	C,	71.70;	H,	3.50;	N,	7.76%.		

7‐amino‐9‐hydroxy‐6‐oxo‐6H‐benzo[c]chromene‐8‐
carbonitrile	 (6):	 FT‐IR	 (KBr,	 cm‐1):	 3370,	 3282,	 3220,	 3079,	
2987,	 2211,	 1705,	 1638,	 1609.	 1H	NMR	 (300	MHz,	DMSO‐d6):	
2.47	 (m,	 3H,	 CH,	 CH2),	 4.22	 (m,	 2H,	 NH2,	 exchangeable	 with	
D2O),	7.43‐7.49	(m,	2H,	H‐2	and	H‐3),	7.72	(dd,	J	=	7.2,	1.2	Hz,	
1H,	 H‐4),	 7.93	 (dd,	 J	 =	 9.0,	 1.2	 Hz,	 1H,	 H‐1),	 9.00,	 (s,	 1H,	

exchangeable	with	D2O),	 9.26	 (s,	 1H,	 exchangeable	with	D2O).	
13C	NMR	(75.46	MHz,	DMSO‐d6):	59.9,	72.1,	116.5,	117.7,	118.5,	
121.0,	124.9,	126.2,	133.2,	151.0,	152.2,	157.1,	164.3,	166.4.	MS	
(EI,	m/z	(%)):	252.5	(M,	5.1).	Anal.	calcd.	for	C14H8N2O3:	66.67;	
H,	3.20;	N,	11.11.	Found:	C,	66.52;	H,	3.00;	N,	11.50%.	

Method	B:	A	solution	of	compound	5	(0.5	g,	0.27	mmol)	in	
ethoxide	 (0.15	 g	 sodium,	 10	 cm3	 absolute	 ethanol),	 and	 then	
ethyl	 cyanoacetate	 (1.0	 cm3)	was	 added	 and	 the	mixture	was	
refluxed	on	water‐bath	for	2	h,	cooled	to	room	temperature	and	
acidified	with	96%	acetic	acid.	The	solid	obtained	was	filtered,	
and	 crystallized	 from	 acetic	 acid	 to	 give	 6	 as	 pall‐yellow	
crystals	(Scheme	2).	M.p.:	237‐238	°C.	Yield:	59%.	
	
2.2.3.	1‐(3‐Cyano‐4‐methyl‐2‐oxoquinolin‐1(2H)‐yl)thiourea	
(7)	
	

A	 mixture	 of	 compound	 5	 (0.25	 g,	 0.13	 mmol)	 and	
thiosemicarbazide	(0.15	g,	0.16	mmol)	in	pyridine	(2	cm3)	was	
refluxed	 for	 5	 h.	 The	 solid	 obtained	 was	 filtered,	 and	
crystallized	from	ethanol	to	give	7	as	orange	crystals	(Scheme	4	
and	5).		

1‐(3‐Cyano‐4‐methyl‐2‐oxoquinolin‐1(2H)‐yl)thiourea	 (7):	
Yield:	 73%.	 M.p.:	 186‐187	 °C.	 FT‐IR	 (KBr,	 cm‐1):	 3368,	 3263,	
3177,	3061,	2923,	2228,	1724,	1601.	1H	NMR	(300	MHz,	DMSO‐
d6):	2.73	(s,	3H,	CH3),	4.47	(s,	2H,	NH2,	exchangeable	with	D2O),	
7.45‐7.50	(m,	2H,	H‐6	and	H‐8),	7.77	(dd,	J	=	7.8,	1.8	Hz,	1H,	H‐
5),	 7.93	 (dd,	 J	 =	 7.8,	 6.3	 Hz,	 1H,	 H‐7),	 8.55	 (s,	 1H,	 NH,	
exchangeable	with	D2O).	13C	NMR	(75.46	MHz,	DMSO‐d6):	18.1,	
101.2,	 114.1,	 116.9,	 118.0,	 120.7,	 125.2,	 127.0,	 135.2,	 152.6,	
156.6,	 163.6.	 MS	 (EI,	 m/z	 (%)):	 256	 (M‐2,	 16.7),	 257	 (M‐1,	
16.7),	 258	 (M,	 13.3).	 Anal.	 calcd.	 for	 C12H10N4OS:	 C,	 55.80;	 H,	
3.90;	N,	 21.69;	 S,	 12.41.	 Found:	 C,	 55.59;	H,	 4.00;	N,	 21.20;	 S,	
12.15%.	



El‐Shaaer	/	European	Journal	of	Chemistry	3	(1)	(2012)	51‐56	 53	
 

	

	
	

Scheme	3	
	
	

	
	

Scheme	4	
	
	

 

	
	

Scheme	5
	
	
2.2.4.	4‐(2‐(6,8‐Dichloro‐4‐oxo‐4H‐chromen‐3‐yl)‐2‐
hydroxyethyl)‐2‐oxo‐2H‐chromene‐3‐carbonitrile	(9)	
	

Method	A:	A	mixture	of	compound	5	(0.5	g,	0.27	mmol)	and	
6,8‐dichloro‐4‐oxo‐4H‐chromene‐3‐carboxaldehyde	(8)	(0.66	g,	
0.27	 mmol)	 in	 dimethylformamide	 (10	 cm3)	 was	 stirred	 at	
room	temperature	(25	°C)	 for	4	h,	 leave	overnight,	 filtered	off	
and	 crystallized	 from	 acetic	 acid	 to	 give	9	 as	 a	white	 crystals	
(Scheme	6).	

4‐(2‐(6,8‐Dichloro‐4‐oxo‐4H‐chromen‐3‐yl)‐2‐hydroxyethyl)‐
2‐oxo‐2H‐chromene‐3‐carbonitrile	 (9):	 Yield:	 60%.	 M.p.:	 293‐
294	 °C.	FT‐IR	 (KBr,	 cm‐1):	3743	 (br),	3060,	2226,	1727,	1652.	
1H	NMR	(500	MHz,	DMSO‐d6):	2.07	(d,	J	=	6.9	Hz,	2H,	CH2),	2.85	
(br.	s,	1H,	CH),	7.39‐7.52	(m,	2H,	Ar‐H),	7.80	(s,	1H,	H‐7),	7.94	
(d,	J	=	8.4	Hz,	1H,	Ar‐H),	8.04	(s,	1H,	H‐5),	8.24	(dd,	J	=	9.95,	6.1	
Hz,	1H,	Ar‐H),	9.08	(s,	1H,	H‐2),	10.05	(s,	1H,	OH).	MS	(EI,	m/z	
(%)):	410	(M‐H2O,	43.3).	Anal.	calcd.	for	C21H11Cl2NO5:	C,	58.90;	
H,	2.59;	N,	3.27.	Found:	C,	58.75;	H,	2.23;	N,	3.56%.	

Method	B:	A	mixture	of	compound	5	(0.5	g,	0.27	mmol)	and	
6,8‐dichloro‐4‐oxo‐4H‐chromene‐3‐carboxaldehyde	(8)	(0.66	g,	
0.27	mmol)	 in	dimethylformamide	 (5	cm3)	was	 refluxed	 for	 5	
min.	The	 solid	 obtained	was	 filtered	 and	purified	 in	 the	 same	
manner	as	in	method	A,	Yield:	69%.	
	
2.2.5.	7‐((6,8‐Dichloro‐4‐oxo‐4H‐chromen‐3‐
yl)methyleneamino)‐9‐hydroxy‐6H‐benzo[c]chromen‐6‐one	
(12)	
	

A	 solution	 of	 compound	5	 (0.5	 g,	 0.27	mmol)	 in	 ethoxide	
(0.15	 g	 sodium,	 10	 cm3	 absolute	 ethanol),	 and	 then	 ethyl	
acetate	 (1.0	 cm3)	was	added	and	 the	mixture	was	 refluxed	on	
water‐bath	for	2	h,	cooled	to	room	temperature	and	filtered	off.	
The	 solid	 obtained	 (0.27	 g)	 was	 mixed	 with	 acetic	 acid	 (2.5	
cm3),	 sodium	 acetate	 (0.3	 g)	 and	 6,8‐dichloro‐4‐oxo‐4H‐
chromene‐3‐carboxaldehyde	 (0.26	 g,	 0.1	 mmol)	 and	 refluxed	
for	1	h,	filtered	off	and	crystallized	from	ethanol	to	give	12	as	a	
pale	orange	crystals	(Scheme	7).	



54	 El‐Shaaer	/	European	Journal	of	Chemistry	3	(1)	(2012)	51‐56	
	

	

	
	

Scheme	6
	

	
7‐((6,8‐Dichloro‐4‐oxo‐4H‐chromen‐3‐yl)methyleneamino)‐

9‐hydroxy‐6H‐benzo[c]chromen‐6‐one	 (12):	 Yield:	 65%.	 M.p.:	
183‐184	 °C.	 FT‐IR	 (KBr,	 cm‐1):	 3431	 (br),	 3075,	 1716,	 1653,	
1602.	 1H	NMR	(500	MHz,	DMSO‐d6):	6.69	 (s,	1H,	 ‐CH=N),	7.25	
(s,	1H,	Ar‐H),	7.37‐7.43	(m,	2H,	Ar‐H),	7.63‐7.72	(m,	3H,	Ar‐H),	
7.96	(s,	1H,	H‐5),	8.02	(s,	1H,	H‐2),	8.16‐8.22	(m,	1H,	Ar‐H),	8.92	
(s,	 1H,	 OH,	 exchangeable	 with	 D2O).	 Anal.	 calcd.	 for	
C23H11Cl2NO5:	C,	61.08;	H,	2.45;	N,	3.10.	Found:	C,	61.42;	H,	2.15;	
N,	2.75%.	
	
2.3.	Antimicrobial	activity	
	

The	newly	 synthesized	 compounds	were	 screened	 against	
Gram‐positive	 bacteria:	 Staphylococcus	 aureus	 (ATCC 25923)	
and	 Bacillus	 subtilis	 (ATCC 6635),	 Gram‐negative	 bacteria:	
Salmonella	 typhimurium	 (ATCC	 14028)	 and	 Escherichia	 coli	
(ATCC	 25922),	 Yeast:	 Candida	 albicans	 (ATCC	 10231)	 and	
Fungus:	 Aspergillus	 fumigatus.	 The	 standardized	 disc‐agar	
diffusion	method	[16]	was	followed	to	determine	the	activity	of	
the	synthesized	compounds	against the	tested	microorganisms.	

The	 tested	 compounds	 were	 dissolved	 in	 dimethyl	
formamide	(DMF)	solvent	and	prepared	in	two	concentrations	
2	 and	 1	 mg/mL.	 The	 antibiotic	 chloramphencol	 was	 used	 as	
standard	 reference	 in	 the	 case	 of	 Gram‐negative	 bacteria,	
Cephalothin	 was	 used	 as	 standard	 reference	 in	 the	 case	 of	
Gram‐positive	 bacteria	 and	 Cycloheximide	 was	 used	 as	
standard	reference	in	the	case	of	yeasts	and	fungi.	
	
3.	Results	and	discussion	
	
3.1.	Chemistry	
	

The	 Claisen	 condensation	 of	 2‐hydroxyacetophenone	
derivatives	with	ethyl	acetate	in	the	presence	of	sodium	metal	
gave	 β‐dicarbonyl	 compound	derivatives	which	were	 cyclized	
under	the	effect	of	conc.	sulfuric	acid	to	give	2‐methylchromone	
derivatives	[15].	

The	 novel	 and	 unexpected	 Claisen	 condensation	 of	 3,5‐
dichloro‐2‐hydroxyacetophenone	 (1)	 with	 excess	 of	 ethyl	
cyanoacetate	 in	 the	 presence	 of	 sodium	metal	 gave	 7‐amino‐
2,4‐dichloro‐9‐hydroxy‐6‐oxo‐6H‐benzo[c]chromene‐8‐carbo‐
nitrile	 (2)	 (Scheme	 1),	 but	 when	 unsubstituted	 2‐
hydroxyacetophenone	 (3)	 reacted	 under	 the	 same	 condition	
gave	 a	mixture	 of	4‐methyl‐2‐oxo‐2H‐chromene‐3‐carbonitrile	
(5)	 (20%)	 and	 7‐amino‐9‐hydroxy‐6‐oxo‐6H‐benzo[c]	
chromene‐8‐carbonitrile	 (6)	 (30%).	 When	 compound	 5	 was	
reacted	 with	 ethyl	 cyanoacetate	 in	 the	 presence	 of	 sodium	
ethoxide	gave	compound	(6)	(59%)	(Scheme	2).	

The	 formation	 of	 compounds	2	 and	6	proceeded	 via	,β‐
unsaturated	ester	derivatives	(4),	which	can	be	cyclized	under	
the	 effect	 of	 heat	 to	 give	 compound	 5,	 further	 Claisen	
condensation	on	active	methyl	group	of	compound	5	 followed	
by	nucleophilic	cyclization	gave	compounds	(2,	6)	(Scheme	3).	

The	 Claisen	 condensation	 of	 3,5‐dichloro‐2‐hydroxy‐
acetophenone	 (1)	 with	 ethyl	 cyanoacetate	 gave	 only	
cyclocondensation	 product	2a	 	2b,	 while	 unsubstituted	 2‐
hydroxyacetophenone	 (3)	 gave	 a	 mixture	 of	 condensation	
product	5	and	cyclocondensation	product	6a	 	6b	due	to	the	
thermodynamic	stabilities	of	2a	 	2b	 than	6a	 	6b	 (Table	
1).	Also,	 theoretical	 thermodynamic	data	 obtained	 from	semi‐
empirical	 AM1‐MO	 calculations	 shows	 that	 structure	 2b	 is	
more	stabilized	than	2a	and	6b	than	6a	(Table	1).	

	
Table	 1.	 Calculated	 heat	 of	 formation	 of	 coumarin	 derivatives	 by	 semi‐
empirical	AM1‐MO	method.	
Compound Heat	of	formation	(kcal/mol)	
2a ‐13.65
2b ‐43.63
5 ‐2.51
6a ‐5.21
6b ‐34.77
9	 ‐72.45	
10	 59.69	

	
The	 presence	 of	 cyano	 group	 in	 a	 3‐position	 of	 coumarin	

moiety	 facilitate	 the	 nucleophilic	 ring	 addition	 on	 C=O	 group	
followed	 by	 ring	 fission	 and	 recyclization,	 so	 the	 reaction	 of	
thiosemicarbazide	 with	 compound	 5	 in	 pyridine	 gave	 1‐(3‐
cyano‐4‐methyl‐2‐oxoquinolin‐1(2H)‐yl)thiourea	 (7)	 (Scheme	
4)	and	the	mechanism	of	formation	of	compound	7	(Scheme	5).	

The	 reaction	 of	 compound	 5	 with	 6,8‐dichloro‐4‐oxo‐4H‐
chromene‐3‐carboxaldehyde	 (8)	 in	 dimethylformamide	 at	
room	 temperature	 (25	 oC)	 and/or	 reflux	 for	 5	 min	 gave	
addition	 product	9	 rather	 than	 the	 condensation	 product	10.	
The	preferable	formation	of	9	rather	than	10	seems	to	be	due	
to	 the	 thermodynamic	 stabilities	 according	 to	 semi‐empirical	
AM1	calculation	data,	where	 the	ΔH°f	 =	 ‐72.45	kcal/mol	 for	9	
which	is	much	more	stabilized	than	ΔH°f	=	59.69	kcal/mol	of	10	
(Table	 1)	 and	 also	 for	 the	 presence	 of	 strong	 hydrogen	 bond	
between	 the	 C=O	 and	 the	 OH	 groups	 of	 chromone	moiety	 of	
compound	9	(Scheme	6).	

The	Claisen	condensation	of	compound	5	with	ethyl	acetate	
in	 the	presence	of	sodium	ethoxide	gave	the	sodium	salt	of	7‐
amino‐9‐hydroxy‐6H‐benzo[c]chromen‐6‐one	 (11)	 as	
intermediate	 product	 which	 upon	 treatment	 with	 8	 in	 acetic	
acid	 and	 sodium	 acetate	 gave	 7‐((6,8‐dichloro‐4‐oxo‐4H‐
chromen‐3‐yl)methyleneamino)‐9‐hydroxy‐6H‐benzo[c]	
chromen‐6‐one	(12)	(Scheme	7).	



El‐Shaaer	/	European	Journal	of	Chemistry	3	(1)	(2012)	51‐56	 55	
 

	

	
	

Scheme	7	
	
	
3.2.	Molecular	orbital	calculations	

	
The	 experimental	 IR	 frequencies	 of	 C=O	 groups	 of	

coumarin	 derivatives	 were	 compared	 with	 theoretical	 bond	
lengths	 of	 C=O	 groups	 which	 were	 obtained	 from	 molecular	
mechanical	calculations	on	the	basis	of	the	semi‐empirical	AM1	
and	Ab	Initio	(STO‐3G)	methods	of	HyperChem	8.03	computer	
program	 after	 geometrical	 optimization	 of	 the	 structures	 for	
compounds	(2‐12)	(Table	2).	

	
Table	2.	 Calculated	 bond	 lengths	 of	 C=O	 of	 coumarin	 derivatives	 by	 semi‐
empirical	 AM1	 and	 Ab	 Initio	 (STO‐3G)	methods	 and	 their	 experimental	 IR	
C=O	values	for	compounds	(2a‐12).	

Compound	
Bond	lengths	of	C=O,	Å	

Experimental					
C=O		(cm‐1)	Semi‐empirical	

(AM1)	
Ab	Initio	
(STO‐3G)	

2a	 1.22484a	 1.21413	 1736
2a	 1.2303b	 1.21421	 1682
5	 1.228a	 1.217	 1723
6a	 1.22623a	 1.21621	 1705
6a	 1.2309b 1.21456	 1680
7	 1.2353c	 1.2208	 1724
9	 1.22853a	 1.21732	 1727	
9	 1.23831b	 1.44367	 1652	
12	 1.22777a	 1.21652	 1716	
12	 1.23908b	 1.45056	 1653
rd	 0.914	 0.987	 ‐	
a	COcoumarin	
b	C=Ochromone	or	cyclic	ketone	
c	C=Oquinolinone		
d	r	=	regression	coefficient.	

	
The	calculated	bond	lengths	of	C=O	groups	(Å)	on	the	basis	

of	 semi‐empirical	 AM1	 method	 are	 linearly	 related	 to	 the	
measured	 IR	 C=O	 groups	 (cm‐1)	 for	 compounds	 (2‐12)	 and	
from	 the	 linear	 relation	 bond	 length	 (C=O)	 =	 1.47‐0.001	C=O,					
r	 =	 0.9140	 except	 (7),	 where	 r	 is	 regression	 coefficient.	 The	
negative	slope	reveals	indirect	proportionality	of	the	calculated	
bond	lengths	with	measured	C=O	values,	which	agreement	with	
Hooke’s	law	and	these	support	the	proposed	structures	for	the	
prepared	compounds.	On	the	other	hand,	when	Ab	Initio	(STO‐
3G)	method	was	used,	 the	 linear	relation	bond	length	(C=O)	=	
1.11+6.25x10‐5	 C=O,	 r	 =	 0.9870	 except	 (2a,	 7,	 9b	 and	 12b)	
which	is	less	efficient	method	than	the	last	method.	

Also,	 the	 calculated	 net	 carbon	 charges	 by	 semi‐empirical	
AM1	 and	 Ab	 Initio	 (STO‐3G)	 methods	 after	 geometrical	
optimization	 were	 compared	 with	 experimental	 13C	 NMR	 	
values	for	compound	5	(Table	3).	

The	 calculated	 net	 carbon	 charges	 on	 the	 basis	 of	 semi‐
empirical	AM1	method	are	linearly	related	to	the	experimental	
13C	 NMR	 (	 in	 ppm)	 for	 compound	 5	 and	 from	 the	 linear	

relation	 charges	 on	 carbon	 atoms	 =	 ‐0.74	 +	 0.005	 13C	 NMR,										
r	 =	 0.9200,	 except	 (C‐2	 and	 C‐10),	 where	 r	 is	 regression	
coefficient.	The	positive	slope	reveals	direct	proportionality	of	
the	 calculated	 net	 carbon	 charges	 with	 measured	 13C	 NMR	 	
values,	which	support	the	proposed	structure	for	compound	5.	
Also,	 when	 Ab	 Initio	 (STO‐3G)	 method	 was	 used,	 the	 linear	
relation	 charges	 on	 carbon	 atoms	 =	 ‐0.25	 +	 0.002	 13C	 NMR,									
r	 =	 0.8340,	 except	 (C‐2	 and	 C‐9),	 which	 less	 agreement	 with	
experimental	 data	 than	 semi‐empirical	 AM1	 method.	 The	
dependence	 of	 13C	 NMR	 shifts	 on	 the	 net	 carbon	 charges	 for	
compound	5	 is	more	pronounced	 than	 that	 found	of	 IR	 (C=O)	
and	 their	 bond	 lengths	 for	 compounds	 (2‐12),	 as	 indicated	
from	slope	values	of	semi‐empirical	AM1	method.	

	
Table	3.	Calculated	net	carbon	charges	by	semi‐empirical	AM1	and	Ab	Initio	
(STO‐3G)	methods	and	their	experimental	13C	NMR	values	for	compound	5.		

Carbon	
number	

Calculated	net	carbon	charges	
Experimental						
13C	NMR		(ppm)	Semi‐empirical	

(AM1)	
Ab	Initio	
(STO‐3G)	

C‐2	 0.350	 0.319	 164.24	
C‐3	 ‐0.125	 ‐0.042	 118.66	
C‐4 0.098 0.066	 153.25
C‐5 ‐0.067 ‐0.048	 135.86
C‐6 ‐0.159 ‐0.070	 117.54
C‐7 ‐0.073 ‐0.043	 127.69
C‐8 ‐0.150 ‐0.075	 125.89
C‐9a ‐0.153 ‐0.036	 101.85
C‐10a 0.118 0.142	 157.32
C‐9	 ‐0.086	 0.073	 114.80	
C‐10	 ‐0.210	 ‐0.189	 18.79	
r*	 0.920	 0.834	 ‐	
*r	=	regression	coefficient.	
	
3.3.	Antimicrobial	activity	
	

The	newly	 synthesized	 compounds	were	 screened	 against	
Gram‐positive	 bacteria:	 Staphylococcus	 aureus	 (ATCC 25923)	
and	 Bacillus	 subtilis	 (ATCC 6635),	 Gram‐negative	 bacteria:	
Salmonella	 typhimurium	 (ATCC	 14028)	 and	 Escherichia	 coli	
(ATCC	 25922),	 Yeast:	 Candida	 albicans	 (ATCC	 10231)	 and	
Fungus:	 Aspergillus	 fumigatus.	 The	 standardized	 disc‐agar	
diffusion	method	[11]	was	followed	to	determine	the	activity	of	
the	synthesized	compounds	against the	tested	microorganisms.	

Compound	 12	 showed	 high	 activities	 against	 Aspergillus	
fumigates	at	concentration	of	2	mg	and	1	mg,	while	 it	showed	
intermediate	activities	against	Bacillus	subtilis,	Escherichia	coli	
and	 Candida	 albicans.	 Compound	 9	 showed	 intermediate	
activities	 against	Bacillus	 subtilis,	Escherichia	 coli	 and	Candida	
albicans	(Table	4).	
	
	



56	 El‐Shaaer	/	European	Journal	of	Chemistry	3	(1)	(2012)	51‐56	
	
	
Table	4.	Antimicrobial	activities	data	of	compounds	2‐12.	

Sample	

Mean	of	zone	diametera,	mm	
Gram‐positive	bacteria	 Gram‐negative	bacteria Yeastsb	

Staphylococcus	
aureus	

(ATCC	25923)	

Bacillus		
subtilis		

(ATCC	6635)	

Salmonella	
typhimurium		
(ATCC	14028)	

Escherichia
coli		

(ATCC	25922)	

Candida		
Albicans	

	(ATCC	10231)	

Aspergillus	
fumigatus	

Conc.g	 2	 1		 2	 1		 2 1	 2 1	 2 1		 2	 1	
2	 4	Ld	 2	L	 3	L	 ‐c	 2	L ‐ 4	L 2	L 2	L ‐	 ‐	 ‐
5	 3	L	 2	L	 3	L	 ‐	 2	L ‐ 2	L ‐ 7	L 4	L	 ‐	 ‐
7	 ‐	 ‐	 6	L	 4	L	 ‐ ‐ 3	L ‐ 9	L 5	L	 2	L	 ‐
6	 ‐	 ‐	 6	L	 4	L	 ‐	 ‐	 3	L	 ‐	 9	L	 5	L	 2	L	 ‐	
9	 8	L	 5	L	 15	Ie	 11	I	 ‐	 ‐	 18	I	 12	I	 18	I	 13	I	 8	L	 5	L	
12	 6	L	 3	L	 13	I	 9	L	 4	L ‐ 17	I 12	I 20	I 17	I	 28	Hf	 20	H
Controlh	 42	 28	 38	 30	 36	 25	 38	 30	 40	 28	 40	 31	

a	Calculate	from	3	values.	
b	Identified	on	the	basis	of	routine	cultural,	morphological	and	microscopical	characteristics.	
c	‐:	No	effect.	 
d	L:	Low	activity	=	Mean	of	zone	diameter	≤	1/3	of	mean	zone	diameter	of	control.	
e	I:	Intermediate	activity	=	Mean	of	zone	diameter	≤	2/3	of	mean	zone	diameter	of	control.	
f	H:	High	activity	=	Mean	of	zone	diameter	>	2/3	of	mean	zone	diameter	of	control.	
g	Concentration,	mg/mL.	
h	Chloramphencol	in	the	case	of	Gram‐positive	bacteria,	Cephalothin in	the	case	of	Gram‐negative	bacteria	and	cycloheximide	in	the	case	of	fungi.	
	
4.	Conclusion	
	

The	 Claisen	 condensation	 of	 2‐hydroxaceophenone	
derivatives	 with	 ethyl	 cyanoacetate	 gave	 condensation	 and	
cyclocondensation	 products	 with	 different	 nucleophilic	
reagents	 and	 depends	 on	 the	 reactivity	 of	 2‐hydroxy‐
acetophenone,	 nucleophilic	 reagents,	 and	 the	 thermodynamic	
stabilities	of	the	products.	Also,	 the	antimicrobial	activities	for	
the	 prepared	 compounds	 indicated	 that	 when	 coumarin	
derivatives	 bear	 a	 chromone	 moiety,	 the	 biological	 activities	
are	increased	compared	with	other	prepared	derivatives.	
	
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