Novel 3-alkanoyl/aroyl/heteroaroyl-2H-chromene-2-thiones: Synthesis and evaluation of their antioxidant activities

Article abstract of DOI:10.1016/j.ejmech.2010.01.070

A facile, convenient and high yielding synthesis of a combinatorial library of 3-alkanoyl/aroyl/heteroaroyl-2H-chromene-2-thiones has been developed by the condensation of easily accessible β-oxodithioesters and salicylaldehyde/substituted 2-hydroxybenzal

Full text of DOI:10.1016/j.ejmech.2010.01.070

European Journal of Medicinal Chemistry 45 (2010) 2250–2257
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Novel 3-alkanoyl/aroyl/heteroaroyl-2H-chromene-2-thiones: Synthesis and
evaluation of their antioxidant activities
,
a
b
Okram Mukherjee Singh a , Nepram Sushuma Devi , Dhanaraj Singh Thokchom ,
*
Gurumayum Jitendra Sharma b
a Department of Chemistry, Manipur University, Canchipur, Imphal 795003, Manipur, India
b Genetics and Radiation Biology Laboratory, Department of Life Sciences, Manipur University, Canchipur, Imphal 795003, Manipur, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A facile, convenient and high yielding synthesis of a combinatorial library of 3-alkanoyl/aroyl/hetero-
Received 3 November 2009
Received in revised form
27 January 2010
Accepted 28 January 2010
Available online 2 February 2010
aroyl-2H-chromene-2-thiones has been developed by the condensation of easily accessible
b-oxodi-
thioesters and salicylaldehyde/substituted 2-hydroxybenzaldehydes under solvent-free conditions. The
assessment of radical scavenging capacity of the compounds towards the stable free radical 2,2-diphenyl-
1-picrylhydrazyl (DPPH) was measured and these compounds were found to scavenge DPPH free radical
efficiently. Five selected compounds were able to protect curcumin from the attack of sulfur free radical
generated by radiolysis of glutathione (GSH). The newly synthesized compounds exhibited profound
antioxidant activities. Five of them rendered comparatively high antioxidant capacity.
Ó 2010 Elsevier Masson SAS. All rights reserved.
Keywords:
Coumarin
Piperidine
b-Oxodithioester
Antioxidant
Free radical
1. Introduction
and exchange resins [20]. Applications of microwaves [21] and ionic
liquids [22] for the synthesis of coumarins have also been reported.
Coumarins constitute an important class of oxygen heterocycles
found abundantly in numerous naturally occurring products,
including edible vegetables and fruits [1]. They are used as additives
in food and cosmetics, optical brightening agents and dispersed
fluorescent and laser dyes [2]. The coumarins display a remarkable
array of biochemical and pharmacological actions, such as anti-
fungal, antihypertensive, antioxidant, anti-inflammatory, and
antimicrobial activity [3–8]. Some coumarins are also reported as
chemotherapeutic agents [9–12].
Knoevenagel condensation is reported recently to proceed with
high selectivity and reactivity in the formation of coumarins over
solid base catalysts [23]. Microwave irradiation accelerates these
reactions several-fold with better yields of the products [24].
However, to the best of our knowledge the new methodologies
in all these reported methods use only the b-ketoesters such as
ethyl acetoacetate or methyl acetoacetate and thus structural
variations are limited to those derived from these esters. As a result,
the structures of known coumarins either in improved yields or by
the application of new techniques are always reported. Recently,
There has been, in recent years, a major rekindling of interest in
the studies on the defensive effects of coumarins as antioxidants
[13,14]. Nishiyama et al. reported the stronger antioxidative activ-
b-oxodithioesters [25] instead of b-ketoesters have been applied in
solvent-free multicomponent reactions for synthesis of dihy-
dropyrimidines [26]. In conjunction with our works related with
the synthesis and biological evaluation of heterocycles [26,27], we
ities of hydrocoumarins than
a-tocopherol for the oxidation of
tetralin and linoleic acid in a homogeneous solution [15].
It is therefore of utmost importance that the synthesis of
coumarin and its derivatives should be achieved by a simple and
effective method. The Pechmann reaction for the synthesis of
have investigated the cyclocondensation of b-oxodithioesters and
2-hydroxy benzaldehyde/substituted salicylaldehydes to yield the
3-alkanoyl/aroyl/heteroaroyl-2H-chromene-2-thiones 5a–o in
good to excellent yields.
coumarin involves the condensation of phenols with
b-ketonic
esters in the presence of variety of acidic condensing agents [16–19]
Furthermore, scavenging activity towards 2,2-diphenyl-1-pic-
rylhydrazyl (DPPH) radical was measured for all the newly
prepared compounds and discussed herein. The antioxidant eval-
uation was supplemented by sulfur free radical reactivity assay
using the technique of radiolysis of glutathione (GSH) and
* Corresponding author. Tel.: þ91 0385 2435307; fax: þ91 0385 2435145.
E-mail address: ok_mukherjee@yahoo.co.in (O.M. Singh).
0223-5234/$ – see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.01.070

O.M. Singh et al. / European Journal of Medicinal Chemistry 45 (2010) 2250–2257
2251
measuring the protective indices of inhibition of curcumin
depletion.
oxodithioesters (3b–g) and different substituted 2-hydrox-
ybenzaldehydes to afford coumarin (5b–o) in good to excellent
yields. The results are summarized in Table 1. The reaction proceeds
smoothly in a short time and the methodology was found equally
facile in all the aryl, alkyl and heteroaryl particularly, 4-pyridyl
2. Results and discussion
substituted
b-oxodithioesters. The functional groups either elec-
2.1. Chemistry
tron withdrawing or electron donating groups at the 3- and 5-
positions were introduced in this coumarin ring systems
successfully.
The syntheses of 3-alkanoyl/aroyl/heteroaroyl-2H-chromene-2-
thiones 5 are summarized in Scheme 1.
b-Oxodithioesters 3a–g are selected as the appropriate synthons
of the present investigation. They are generally prepared by
reacting any active methylene compound with CS2 in the presence
of a suitable base followed by alkylation with either dimethylsulfate
or methyl iodide. However, this method gives poor yields and also
mixtures of dithioesters and ketene dithioacetals. Thus improved
2.2. Antioxidant assay
2.2.1. DPPH free radical scavenging assay
The use of stable free radical DPPHꢁ in evaluating the free radical
scavenging activity of extracts from plants, food materials and
single compounds [28] is one of the most frequently used methods.
The assay is based on the assessment of scavenging capacity of
antioxidants towards the stable free radical DPPHꢁ which is indi-
cated by the bleaching of purple coloured methanol solution of
DPPHꢁ to the corresponding yellow coloured hydrazine [29] either
by hydrogen atom- or electron donation [30].
yields of b-oxodithioesters are obtained by treating active methy-
lene compounds 1a–g with (S,S)-dimethyl trithiocarbonate.
Subsequently, these intermediates were treated for Knoevenagel
type condensations with ortho hydroxyl substituted benzaldehydes
to afford the desired 3-alkanoyl/aroyl/heteroaroyl-2H-chromene-
2-thiones 5 in good to excellent yields (Scheme 1).
Several catalytic and noncatalytic conditions were evaluated for
this cyclocondensation reaction. We found the best yields of the
desired coumarins by using piperidine as the base and no other
organic solvent was used as a medium of the reaction.
Previously we have carried out the coumarin synthesis by
heating b-oxodithioester 3a (2.5 mmol) with 2-hydroxy benzalde-
hyde 4a (2.5 mmol) at 90 ꢀC for 45 min in the presence of piperi-
dine (2.5 mmol). After completion of the reaction (monitored by
TLC), water was added, and the product was extracted with ethyl
acetate. After the organic layer was dried (Na2SO4) and evaporated,
the residue was recrystallized from ethyl acetate and hexane to give
the desired coumarin product 5a as yellow crystals in 93% yield. The
formation of compound 5a was confirmed by the spectral and
analytical data.
The use of DPPH in antioxidant assays is well documented
probably because of its rapid reaction and reliability. Initially, free
radical scavenging activities of the newly prepared compounds 5a–
o were evaluated based on the DPPH decoloration range. Three
compounds 5a, 5f and 5m were omitted in the discussion part for
clarity as their antioxidant activity from the initial measurements
of these compounds using DPPH was found almost negligible
(below 5%).
Thus the wide range of activities of the 12 tested compounds
excluding 5a, 5f and 5m were found from 6.41% to 92.81% at
a concentration of 250 mg/mL. Compound 5l was found to be the
most potent antioxidant with the least values of IC50 (inhibition
concentration) and the least active among the series being
compound 5b with the largest values of IC50 (Table 2 and Fig. 1).
It is noteworthy to mention that using different solvents such as
DMSO, DMF, THF and acetonitrile did not improve the yields and
thus we have optimized the reaction condition at 90 ꢀC for 2 h
under solvent-free condition. After optimizing the reaction condi-
The IC50 values were found to vary from 31.20
1579.25 g/mL showing awide rangeof variations in the reactivityof
the samples. Compound 5l having the least IC50 value of 31.20 g/mL
mg/mL to
m
m
and the highest corresponding value of AEAC (ascorbic acid equiv-
tion, the same process was successfully extended to different
b
-
alent antioxidant capacity), was followed by 5e, 5i, 5g and 5j in the
O
S
O
S
NaH
C6H6, 60-70oC
+
R1
CH3
MeS
R1
SMe
SMe
3
1
2
R2
CHO
OH
Piperidine
90oC
R3
4
O
R2
R1
O
S
R3
5
Scheme 1. Synthesis of 3-alkanoyl/aroyl/heteroaroyl-2H-chromene-2-thiones 5a–o.

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O.M. Singh et al. / European Journal of Medicinal Chemistry 45 (2010) 2250–2257
Table 1
Piperidine catalyzed preparation of coumarins under solvent-free conditions.a
O
S
R2
CHO
OH
R2
O
S
R1
Piperidine
90oC
+
R1
SMe
O
R3
R3
3a-g
4a-c
5a-o
Entry
1.
R1
R2
R3
Product (5)
Yieldb (%)
O
H
H
H
H
H
H
93
O
S
5a
O
S
2.
3.
95
90
H3CO
O
OCH
3 5b
O
Cl
O
S
Cl
5c
O
4.
5.
6.
H
H
H
H
H
H
95
91
85
H3C
O
S
5d
O
S
CH3
O
5e
O
N
N
O
S
5f
O
7.
H
OCH3
93
O
S
OCH3
5g
O
8.
H
OCH3
91
H3CO
O
S
OCH3
OCH3
5h

O.M. Singh et al. / European Journal of Medicinal Chemistry 45 (2010) 2250–2257
2253
Table 1 (continued)
Entry
R1
R2
R3
Product (5)
Yieldb (%)
O
9.
H
OCH3
95
O
S
Cl
Cl
OCH3
5i
O
S
10.
H
OCH3
92
H3C
O
OCH3
5j
O
11.
CH3
H
OCH3
89
O
S
OCH3
5k
O
S
S
12.
H
OCH3
87
S
O
OCH3
5l
O
Br
13.
14.
Br
H
91
O
S
5m
O
Br
Br
Br
H
H
85
89
H3CO
O
S
OCH3
5n
O
Br
15.
Cl
O
S
Cl
5o
a
Reaction conditions: 3 (2.5 mmol), 4 (2.5 mmol), piperidine (2.5 mmol), 90 ꢀC, 1.0–2.0 h.
Isolated yield.
b
activity test. Compound 5b with the IC50 value of 1579.25
mg/mL
Fig. 2. The reaction solutions supplemented with five compounds
5e, 5g, 5i, 5j and 5l, which were selected through assessment of IC50
and AEAC values from the DPPH free radical scavenging assay
exhibited substantial reduction in depletion of curcumin with
variations. Reaction solutions supplemented with compounds 5l
showed the lowest DPPHꢁ scavenging activity (Table 2). The wide
variations in free radical scavenging activities may be due to the
variations in the proton–electron transfer by the compounds due to
difference in their structures and stability.
and 5e at a concentration of 10
by 2.41 M and 3.27 M respectively at a dose of 50 Gy. The samples
showed varying protective indices (Table 3). Calculations were
done using a concentration of 10 g/mL for the five samples. A
relative assessment of the PIs (protective indices) of the five tested
mg/mL inhibited curcumin depletion
2.2.2. Sulfur free radical reactivity assay
m
m
Increase in the depletion of curcumin content with increase in
dose of
g
-radiation was observed in the control reaction solutions.
m
Curcumin depletion by free radicals (GSꢁ) in the reactions is given in

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O.M. Singh et al. / European Journal of Medicinal Chemistry 45 (2010) 2250–2257
Table 2
and AEAC values may be attributes of variation in molecular
structures and nature of the compounds. Interestingly, the 2-acetyl
thiophene substituted compound 5l was found to be the most
potent antioxidant in the series of the newly prepared compounds.
Free radical scavenging capacities of coumarin derivatives measured in DPPH assay.
Sl. no.
Sample
Mean IC50 ꢃ SE (
mg/mL)
1
2
3
4
5
6
7
8
5b
5c
5d
5e
5g
5h
5i
5j
5k
5l
1579.25 ꢃ 0.64
537.58 ꢃ 0.79
551.57 ꢃ 0.80
35.01 ꢃ 0.56
98.72 ꢃ 0.97a
603.43 ꢃ 0.57
95.57 ꢃ 0.91a
233.71 ꢃ 0.70
358.67 ꢃ 0.76
31.20 ꢃ 0.40
1112.83 ꢃ 0.90
850.41 ꢃ 0.69
3. Conclusion
In conclusion, we have successfully demonstrated the synthesis
and antioxidant evaluation of 3-alkanoyl/aroyl/heteroaroyl-2H-
chromene-2-thiones.
b-Oxodithioesters have been utilized as
9
reactive intermediates to afford the desired coumarins by treating
them with salicylaldehyde/substituted 2-hydroxybenzaldehydes in
presence of piperidine under solvent-free conditions. The antioxi-
dant activities of the 12 newly prepared compounds were
measured using DPPH and we found them to be efficient DPPH free
radical scavengers. Further, five selected compounds were found as
protectors of curcumin from the attack of sulfur free radical
generated by radiolysis of glutathione (GSH).
10
11
12
5n
5o
a
Values of IC50 are expressed as means ꢃ SE (n ¼ 3). Values followed by same
letters do not differ significantly at P > 0.05 as measured by Tukey HSD test.
compounds at 100 Gy revealed that the activity order was as
5l > 5e > 5i > 5g > 5j (Fig. 3). However, the order of PIs changes
with variation in radiation dose, and decrease with increasing
radiation dose.
4. Experimental
Aqueous solution of glutathione (GSH) generates sulfur free
radicals (GSꢁ) when irradiated with -radiation. The GSꢁ free radical
g
4.1. General
resulting from the irradiation of GSH oxidizes the chrome yellow
compound curcumin present in the reaction solution and depletes
its colour. Inhibition of curcumin depletion by substitution of 60%
methanol solution of the selected compounds suggests that they
possess certain entities which contribute to free radical scavenging
activities and antioxidant properties. Substitution of the reaction
solutions with the samples protect the curcumin molecules in the
solution from getting depleted due to GSꢁ free radicals even at
a high dose of 250 Gy.
The PI values decrease as the dose increases as the curcumin
molecules are not able to compete with the sulfur free radicals (GSꢁ)
produced after a certain threshold dose. More damages are caused
to the curcumin molecules as the generation of free radicals
increase with increasing dose. The compounds 5l and 5e showed
comparatively good response in terms of DPPH stable radical
scavenging and inhibition of curcumin depletion by GSꢁ radicals as
compared to the other samples (Table 3). The variation in PI, IC50
Melting points are uncorrected and were determined in capil-
lary tubes on an apparatus containing silicon oil. The IR spectra
were recorded on a Perkin Elmer 983 spectrometer in KBr pellets
with absorption given in cmꢂ1 1H and 13C NMR spectra were
.
recorded respectively on a Varian EM-390 (300 MHz and 75.5 MHz)
spectrometer. The chemical shifts ( ppm) and the coupling
d
constants (Hz) are reported in the standard fashion with reference
to internal tetramethyl silane (TMS). The MS spectra were recorded
on a Jeol JMSD-300 spectrometer. Elemental analyses were per-
formed on a Carlo Erba’s108 microanalyzer.
4.2. General procedure for the synthesis of b-oxodithioesters
(3a–g) [25]
NaH (0.05 mol) was added slowly to a solution of the appro-
priate ketone (1; 0.025 mol) in dry benzene (50 mL) followed by
Fig. 1. Variations in IC50 of 12 selected compounds.

O.M. Singh et al. / European Journal of Medicinal Chemistry 45 (2010) 2250–2257
2255
Protection of Curcumin Depletion by Coumarin Derivatives
20
15
10
5
Curcumin + GSH
5e
5g
5i
5j
5l
0
0 Gy
50 Gy
100 Gy
Dose
Fig. 2. Depletion of curcumin in control caused by sulfur free radical (GSꢁ) and its protection by 60% methanol solutions of the selected compounds.
200 Gy
250 Gy
the addition of dimethyl trithiocarbonate (2; 0.027 mol) dropwise
over a period of 1–2 h. The mixture was stirred for 30 min at room
temperature and then refluxed for 2 h. The mixture was allowed to
cool, poured into ice-cold water (250 mL). The aqueous layer was
separated, washed with benzene (200 mL), acidified with 3 N
hydrochloric acid or 20% acetic acid, and extracted with chloroform
(2 ꢄ 150 mL). The extract was dried with sodium sulfate and
evaporated to give the products 3a–g. Purification was done by
column chromatography on silica gel using hexane as eluent.
Spectral data of the unreported compound 3f is given below:
Methyl-3-oxo-3-(pyridine-4-yl)propanedithioate (3f): Yellow
After the organic layer was dried (Na2SO4) and evaporated, the
residue was recrystallized by ethyl acetate and hexane to products
5a–o. In cases where further purification was required, the crude
products were subjected to column chromatography on SiO2, using
increasing amounts of ethyl acetate in hexanes as eluent.
3-Benzoyl-2H-chromene-2-thione (5a). Yellow crystals. m.p. 170–
171 ꢀC. 1H NMR (300 MHz, CDCl3,
d
ppm): 7.37–7.42 (m, 1H), 7.45–
7.56 (m, 3H), 7.59–7.66 (m, 3H), 7.68–7.71 (m, 1H), 7.94–7.96 (m,
2H); 13C NMR (75.5 MHz, CDCl3,
ppm): 116.7, 119.9, 125.9, 128.6,
128.7, 129.6, 133.2, 133.6, 133.9, 135.6, 139.1, 157, 192.3, 193.6; IR
d
(KBr) (n
max, cmꢂ1): 1246, 1604, 1662, 3032, 3052; MS m/z 266
crystals. m.p. 71–72 ꢀC. 1H NMR (300 MHz, CDCl3,
d
ppm): 2.7 (s,
(Mþ). Anal. Calcd. for C16H10O2S: C, 72.16; H, 3.78; S,12.04. Found: C,
72.10; H, 3.72; S, 11.84.
3H); 6.9 (s, 1Holefin); 7.9 (d, J ¼ 6 Hz, 2H); 8.3 (d, J ¼ 6.3 Hz, 2H); 15.0
(s, 1H, OH); 13C NMR (75.5 MHz, CDCl3,
120.5, 145.8, 156.5, 191.9, 220.5; IR (KBr) (
d
ppm): 22.5, 65.1, 119.9,
max, cmꢂ1): 1263, 1585;
3-(4-Methoxybenzoyl)-2H-chromene-2-thione (5b). Yellow crys-
n
tals. m.p. 187–189 ꢀC. 1H NMR (300 MHz, CDCl3,
d ppm): 3.85 (s,
MS: m/z ¼ 211 (Mþ). Anal. Calcd. for C9H9ONS2: C, 51.16; H, 4.29; S,
3H), 6.93 (d, J ¼ 8.7 Hz, 2H), 7.36–7.43 (m, 1H), 7.53–7.71 (m, 4H),
30.35. Found: C, 51.15; H, 4.27; S, 30.37.
7.89 (d, J ¼ 8.7 Hz, 2H); 13C NMR (75.5 MHz, CDCl3,
d ppm): 55.5,
114, 116.7, 120, 125.9, 128.4, 128.6, 132.2, 133, 133.2, 139.5, 157, 164.3,
190.8, 193.8; IR (KBr) (n
max, cmꢂ1): 1242, 1597, 1654, 3018, 3055;
4.3. General procedure for the synthesis of 2H-chromene-2-thiones
(5a–o)
Relative protective indices of Coumarin derivatives at 100 Gy
1000
100
10
Salicyldehyde/substituted salicyldehyde (2.5 mmol), dithioester
(2.5 mmol), and piperidine (10 mol %) were heated at 90 ꢀC with
stirring for 1–2 h. Then the progress of the reaction was monitored
by thin layer chromatography. After completion of the reaction,
water was added, and the product was extracted with ethyl acetate.
Table 3
Ascorbic acid equivalent antioxidant capacity (AEAC) and protective indices (PI) of
the five effective coumarin derivatives.
Sl. No.
Samples
AEAC (mg/g)
PI (%) at 100 Gy
1
2
3
4
5
5e
5g
5i
5j
5l
110.78 ꢃ 0.14
38.70 ꢃ 0.38a
39.98 ꢃ 0.37a
16.38 ꢃ 0.30
121.01 ꢃ 0.10
45.83 ꢃ 0.03
10.94 ꢃ 0.19
25.23 ꢃ 0.04
3.65 ꢃ 0.09
56.93 ꢃ 0.02
1
5j
5g
5i
5e
5l
a
Values of AEAC and PI are expressed as means ꢃ SE (n ¼ 3). Values followed by
same letters in the same column do not differ significantly at P > 0.05 as measured
by Tukey HSD test.
Fig. 3. Relative protective indices (PI) of best five effective compounds against GSꢁ free
radical induced curcumin depletion at 100 Gy.

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O.M. Singh et al. / European Journal of Medicinal Chemistry 45 (2010) 2250–2257
MS: m/z ¼ 296 (Mþ). Anal. Calcd. for C17H12O3S: C, 68.90; H, 4.08; S,
10.82. Found: C, 68.88; H, 3.98; S, 10.79.
3-Acetyl-8-methoxy-2H-chromene-2-thione (5k). Yellow crystals.
m.p. 145–146 ꢀC. 1H NMR (300 MHz, CDCl3,
ppm): 2.66 (s, 3H),
3.93 (s, 3H), 7.07–7.23 (m, 3H), 7.66 (s, 1H); 13C NMR (75.5 MHz,
CDCl3, ppm): 30.0, 56.3, 114.9, 120.1, 120.5, 125.8, 135.4, 140.3,
d
3-(4-Chlorobenzoyl)-2H-chromene-2-thione (5c). Yellow crystals.
m.p. 185–186 ꢀC. 1H NMR (300 MHz, CDCl3,
d
ppm): 7.31–7.35 (m,
1H), 7.42 (d, J ¼ 8.4 Hz, 2H), 7.51 (m, 3H), 7.59–7.66 (m, 3H), 7.68–
7.71 (m, 1H), 7.94–7.96 (m, 2H); 13C NMR (75.5 MHz, CDCl3,
ppm):
116.7, 119.9, 125.9, 128.7, 129.6, 133.2, 133.6, 133.9, 135.6, 139.1, 157,
d
146.6, 147.1, 192.6, 199.4; IR (KBr) (n
max, cmꢂ1) 1276, 1602, 1693,
d
3093; MS m/z ¼ 234 (Mþ). Anal. Calcd. for C12H10O3S: C, 61.52; H,
4.30; S, 13.69. Found: C, 61.55; H, 4.31; S, 13.65.
192.3, 193.6; IR (KBr) (
n
max, cmꢂ1): 1234, 1604, 1660, 3036, 3091;
3(2-Acetylthiophene)-8-methoxy-2H-chromene-2-thione
(5l).
ppm):
MS: m/z ¼ 300.5 (Mþ). Anal. Calcd. for C16H19O2SCl: C, 63.90; H,
Yellow crystals. m.p. 165–167 ꢀC. 1H NMR (300 MHz, CDCl3,
d
3.02; S, 10.66. Found: C, 63.88; H, 3.01; S, 10.59.
3.83 (s, 3H), 6.74–6.79 (m, 2H), 7.06–7.09 (t, 1H), 7.25 (s, 1H), 7.40–
3-(4-Methylbenzoyl)-2H-chromene-2-thione (5d). Yellow crys-
7.42 (m, 1H), 7.63 (d, J ¼ 3.9 Hz,1H), 7.83 (d, J ¼ 2.7 Hz,1H); 13C NMR
tals. m.p. 172–173 ꢀC. 1H NMR (300 MHz, CDCl3,
d
ppm): 2.43 (s,
(75.5 MHz, CDCl3, d ppm): 56.1, 111.5, 119.7, 120.2, 121.2, 125.1, 128.1,
3H), 7.26 (d, J ¼ 8.1 Hz, 2H), 7.36–7.41 (m, 1H), 7.58–7.60 (m, 4H),
134.7, 135.5, 138.7, 143.3, 144.7, 146.4, 185.6, 192.8; MS m/z ¼ 302
(Mþ). Anal. Calcd. for C15H10O3S2: C, 59.58; H, 3.33; S, 21.21. Found:
C, 59.55; H, 3.31; S, 21.18.
7.83 (d, J ¼ 8.4 Hz, 2H); 13C NMR (75.5 MHz, CDCl3,
d ppm): 21.8,
116.6, 119.9, 125.6, 125.8, 128.5, 129.5, 129.8, 133.1, 139.3, 145.1, 157,
191.9, 193.7; IR (KBr) (
n
max, cmꢂ1): 1238, 1604, 1666, 3043, 3059;
3-Benzoyl-6-bromo-2H-chromene-2-thione (5m). Yellow crystals.
MS: m/z ¼ 280 (Mþ). Anal. Calcd. for C17H12O2S: C, 72.83; H, 4.31; S,
m.p. 180–181 ꢀC. 1H NMR (300 MHz, CDCl3,
d ppm): 7.19 (s, 1H),
11.44. Found: C, 72.78; H, 4.28; S, 11.40.
7.33–7.43 (m, 3H), 7.53–7.56 (m, 1H), 7.64–7.69 (m, 2H), 7.85 (d,
3-Acetyl-2H-chromene-2-thione (5e). Yellow crystals. m.p. 178–
J ¼ 5.4 Hz, 2H); 13C NMR (75.5 MHz, CDCl3,
d ppm): 118.3, 118.6,
121.4,128.8,129.6,130.6, 131.7,134.2,135.4,135.9,140.1,155.8,191.8,
179 ꢀC. 1H NMR (300 MHz, CDCl3,
d
ppm): 2.43 (s, 3H), 7.36–7.41 (m,
2H), 7.58–7.60 (m, 3H); 13C NMR (75.5 MHz, CDCl3,
d
ppm): 20.5,
192.8; IR (KBr) (
n
max, cmꢂ1): 1234,1596,1662, 3064; MS m/z ¼ 345
113.9, 119.6, 128.4, 128.6, 129.8, 131.1, 134.5, 156.3, 191.7, 193.7; MS:
m/z ¼ 204 (Mþ). Anal. Calcd. for C11H8O2S: C, 64.69; H, 3.95; S,15.70.
Found: C, 64.78; H, 4.08; S, 15.70.
(Mþ). Anal. Calcd. for C16H9O2SBr: C, 55.67; H, 2.63; S, 9.29. Found:
C, 55.69; H, 2.61; S, 9.31.
3-(4-Methoxybenzoyl)-6-bromo-2H-chromene-2-thione
(5n).
ppm):
3-(4-Acetylpyridine)-2H-chromene-2-thione (5f). Yellow crystals.
Yellow crystals. m.p. 208–209 ꢀC. 1H NMR (300 MHz, CDCl3,
d
m.p. 179–181 ꢀC. 1H NMR (300 MHz, CDCl3,
d
ppm): 7.66–7.80 (m,
4H), 7.99 (d, J ¼ 6 Hz, 1H), 8.18 (d, J ¼ 6.6 Hz, 1H), 8.29–8.35 (m, 1H),
8.51 (s, 1H), 8.81–8.85 (m, 1H); 13C NMR (75.5 MHz, CDCl3,
ppm):
3.8 (s, 3H), 6.87 (d, J ¼ 6.6 Hz, 2H), 7.33 (d, J ¼ 6.6 Hz, 1H), 7.40 (s,
1H), 7.26–7.67 (m, 2H), 7.83 (d, J ¼ 6.6 Hz, 2H); 13C NMR (75.5 MHz,
d
CDCl3, d ppm): 55.6, 114.2, 118.3, 118.5, 121.5, 128.3, 130.5, 131.3,
115.1, 116.5, 121.7, 122.0, 127.3, 129.3, 129.4, 130.7, 135.7, 142.3, 150.9,
192.2, 192.2; MS: m/z ¼ 267 (Mþ). Anal. Calcd. for C15H9NO2S: C,
67.40; H, 3.39; N, 5.24; S, 12.00. Found: C, 67.37; H, 3.37; N, 5.29; S,
11.99.
132.2, 135.7, 140.4, 155.7, 164.5, 190.3, 193.0; IR (KBr) (n
max, cmꢂ1):
1236, 1593, 1643, 3051; MS m/z ¼ 375 (Mþ). Anal. Calcd. for
C17H11O3SBr: C, 54.41; H, 2.95; S, 8.55. Found: C, 54.45; H, 2.93; S,
8.57.
3-Benzoyl-8-methoxy-2H-chromene-2-thione (5g). Yellow crys-
3-(Chlorobenzoyl)-6-bromo-2H-chromene-2-thione (5o). Yellow
tals. m.p. 142–143 ꢀC .1H NMR (300 MHz, CDCl3,
d
ppm): 4.03 (s,
crystals. m.p. 195–196 ꢀC. 1H NMR (300 MHz, CDCl3,
d
ppm): 7.41–
7.55 (m, 3H), 7.73–7.87 (m, 5H); 13C NMR (75.5 MHz, CDCl3,
ppm):
118.4, 118.7, 121.3, 129.1, 129.2, 130.7, 130.9, 132.1, 133.8, 136.1, 139.7,
3H), 7.14–7.20 (m, 2H), 7.27–7.34 (m, 1H), 7.45–7.49 (m, 2H), 7.59–
d
7.62 (m, 2H), 7.94 (d, J ¼ 5.7 Hz, 2H); 13C NMR (75.5 MHz, CDCl3,
d
ppm) 56.3, 114.6, 119.6, 120.6, 125.8, 128.7, 129.6, 133.6, 133.9,
155.8, 190.6, 192.7; IR (KBr) (n
max, cmꢂ1): 1234, 1587, 1662, 3053;
139.5, 146.8, 147.0, 192.4, 192.9; IR (KBr) (
n
max, cmꢂ1): 1274, 1602,
MS m/z ¼ 377 (Mþ). Anal. Calcd. for C16H8O2SClBr: C, 50.62; H, 2.12;
1664, 3951; MS: m/z ¼ 298 (Mþ). Anal. Calcd. for C17H14O3S: C,
68.44; H, 4.73; S, 10.75. Found: C, 68.45; H, 4.77; S, 10.72.
3(4-Methoxybenzoyl)-8-methoxy-2H-chromene-2-thione(5h).
S, 8.45. Found: C, 50.67; H, 2.09; S, 8.43.
4.4. Antioxidant assay
Yellow crystals. m.p. 140–142 ꢀC. 1H NMR (300 MHz, CDCl3,
d ppm):
3.80 (s, 3H), 3.92 (s, 3H), 6.85 (d, J ¼ 6.6 Hz, 2H), 7.04–7.11 (m, 2H),
4.4.1. Materials and methods
7.19–7.24 (m, 1H), 7.48 (s, 1H), 7.83 (d, J ¼ 6.6 Hz, 2H); 13C NMR
4.4.1.1. DPPH free radical scavenging assay. Methanol solution of
2,2-diphenyl-1-picrylhydrazyl (DPPHꢁ) was used as a reagent for
the spectrophotometric assay with modifications [31–33]. The
reaction mixture in triplicates consisted of 100
lute methanol with different concentrations (0–1000
compounds. A negative control with the same DPPH concentration
in methanol without sample was used. Absorbance was read
against a blank at 515 nm after incubation of the reaction mixtures
for 30 min in dark at room temperature. Percentage decoloration of
DPPH was determined by comparison with methanol treated
control and IC50 was calculated in mg/mL. Ascorbic acid equivalent
antioxidant capacity (AEAC) in mg/g was calculated for relatively
effective compounds from the IC50 as
(75.5 MHz, CDCl3, d ppm) 55.6, 56.3, 114.1, 114.42, 119.5,120.7, 125.8,
128.6, 132.2, 133.3, 139.8, 146.8, 146.9, 164.3, 190.9, 193.0; IR (KBr) (
n
max, cmꢂ1): 1274,1593,1651, 3051; MS m/z ¼ 326 (Mþ). Anal. Calcd.
for C18H14O4S: C, 66.24; H, 4.32; S, 9.82. Found: C, 66.25; H, 4.31; S,
9.85.
m
M DPPH in abso-
mg/mL) of the
3(4-Chlorobenzoyl)-8-methoxy-2H-chromene-2-thione
(5i).
ppm):
Yellow crystals. m.p. 175–176 ꢀC. 1H NMR (300 MHz, CDCl3,
d
3.95 (s, 3H), 7.07–7.13 (m, 2H), 7.19–7.26 (m, 1H), 7.37 (d, J ¼ 5.4 Hz,
2H), 7.54 (s, 1H), 7.78 (d, J ¼ 6.3 Hz, 2H); 13C NMR (75.5 MHz, CDCl3,
d
ppm): 56.3, 114.7, 119.7, 120.6, 125.9, 129.1, 130.9, 134.1, 134.1,
139.0, 140.3, 146.8, 147.1, 191.2, 192.7; MS m/z ¼ 330 (Mþ). Anal.
Calcd. for C17H11O3SCl: C, 61.73; H, 3.35; S, 9.69. Found: C, 61.75; H,
3.31; S, 9.65.
AEACðmg Ascorbic acid=g SampleÞ
¼ IC50 ðascorbateÞ=IC50 ðsampleÞ ꢄ 1000:
3(4-Methylbenzoyl)-8-methoxy-2H-chromene-2-thione
(5j).
ppm):
Yellow crystals. m.p. 163–165 ꢀC. 1H NMR (300 MHz, CDCl3,
d
2.43 (s, 3H), 4.03 (s, 3H), 7.13–7.32 (m, 5H), 7.57 (s, 1H), 7.84 (d,
J ¼ 6.3 Hz, 2H); 13C NMR (75.5 MHz, CDCl3,
d
ppm): 21.8, 56.3, 114.5,
The IC50 of ascorbic acid used for calculation of AEAC was
3.82 mg/mL.
119.5, 120.7, 125.7, 129.5, 129.8, 133.2, 133.4, 139.7, 145.1, 146.8,
192.0, 192.9; IR (KBr) (n
max, cmꢂ1): 1276, 1604, 1658, 3058; MS m/
z ¼ 312 (Mþ). Anal. Calcd. for C18H16O3S: C, 69.21; H, 5.16; S, 10.26.
4.4.1.2. Sulfur free radical reactivity assay. Glutathione (GSH) was
used for in vitro generation of sulfur free radical (GSꢁ) by
Found: C, 69.25; H, 5.13; S, 10.29.
g-

O.M. Singh et al. / European Journal of Medicinal Chemistry 45 (2010) 2250–2257
2257
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reactivity with the GSꢁ radicals [34,35]. The reaction solution con-
tained 20
methanol [36]. Various concentrations of samples (5–20
were added to the reaction solution and the final volume was
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ꢂwith sampleÞ=initial curcumin concentrationꢅ ꢄ 100:
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