chapter-3 synthesis, characterization and cytotoxicity...
TRANSCRIPT
Chapter-3
Synthesis, characterization and cytotoxicity
evaluation of nimesulide based
new glycolamide esters
1. Poster at International Conference on “Recent Advances in Drug
Discovery’’ organized by Kakatiya University, Warangal. Oct 22-24,
2008, Pg. No-69.
Chapter-3
106
3.1 INTRODUCTION
The glycolamide ester moiety has been found in many
pharmaceutically important prodrug molecules. Esters of
2-hydroxyacetamide (1) are known as glycolamide esters 2.
ArO
O
NR
R1
O
HO
O
NH2
Unsubstituted glycolamide = R, R1 = H
Monosubstituted glycolamide = R=H, R1=alkyl/aryl
Disubstituted glycolamide = R, R1 = alkyl/aryl
1 2
N,N-Disubstituted glycolamide esters have earlier been reported as
potentially useful biolabile carrier linked prodrug type for carboxylic
acids mainly for NSAIDs.1
Bundgaard and Nielsen have discovered that glycolamides are cleaved
with remarkable speed in human plasma as compared to methyl or ethyl
esters.2 The study was carried out on a series of benzoate esters of
various N-substituted glycolamides 3.
O
O
NR1
R2
O
3
A series of glycolamide ester prodrugs of 6-methoxy-2-napthylacetic
acid3 4 (metabolite of NSAID nabumetone) and 4-biphenyl acetic acid 5
(metabolite of NSAID fenbufen) were synthesized by masking the free
carboxyl group, mainly responsible for gastric damage.4
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107
OMe
O
O
N
O
R
R1
O
O
N
O
R
R1
Ph
4 5
Similarly glycolamide esters of aspirin5 6, ibuprofen6 7, niflumic
acid7 8, scutellarin8 9 as biolabile prodrugs of carboxylic acid agents were
disclosed in the literature.
OCOMe
O
O
NRR1
O
R1RN
O
O
O
Me
Me
Me
6 7
N
HN CF3
O
O
NRR1
O
OHOHO
OH
O
OO
O
OH
OOH
HO
R'RNO
8 9
Apart from the application of glycolamide esters as potent prodrugs,
they have multiple uses.
Glycolamide ester analogues of indomethacin were synthesized and
tested for their cyclooxygenase (COX-1 and COX-2) inhibition properties
in vitro by Smriti et al.9 Compound 10 displayed good anti-inflammatory
activity in vivo.
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NMe
Me
O
O
O
N
O
O
Cl
OMe
O
O
N
O
R
R1
Me
10 11
A series of glycolamide esters of naproxen 11 were synthesized by
Nalini et al.10 The prepared derivatives showed significant anti-
inflammatory, anticonvulsants and reduced ulcerogenic activity when
compared with naproxen.
N-benzhydryl glycolamide esters are used as carboxyl protecting
groups in peptide synthesis.11 Nipecotamide glycolamide esters are used
in the treatment of platelet mediated thrombosis disorder.12 Glycolamide
esters of ibuprofen were found to have comparable anti-inflammatory
and analgesic activity as parent ibuprofen.6 Colfenamate (12)
(carbamoylmethyl 2-(3-(trifluoromethyl)phenylamino)benzoate) having
analgesic, anti-inflammatory activity also belongs to the class of
glycolamide ester.13
F3CHN
O ONH2
O
12
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109
3.2 PRIOR ART ON SYNTHESIS OF GLYCOLAMIDE ESTERS Retrosynthesis of glycolamide ester (retrosynthetic route a & b)
indicate that the synthetic approach may consist of two parts: (i)
construction of amide bond and (ii) development of ester bond
irrespective of order (figure 3.1).
R O
O
NH
O
R'
ab
R O
O
O
C---N
-NH
R'
R'NH2R O
O
NH
O
R'
+RCOOCH2COCl
+
RCOOCH2COOHRCOOCH2CONH2ClCH2CONH2+RCOOH
(a)+
R O
O
NH
O
R'
C---O RCOOH
+
ClCH2CONHR' ClCH2COCl
(b)
+ R'NH2
Figure 3.1: Retrosynthetic pathway of glycolamide ester
Literature search revealed that reports are available for the synthesis
of glycolamide esters based on both strategies.
An overview on selected methods for the synthesis of glycolamide
esters.
A) Construction of ester followed by amide based on retrosynthetic route
a.2
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110
Gycolamide esters were synthesized by reaction of benzoyloxyacetyl
chloride (obtained by reaction between benzoyl glycolic acid and thionyl
chloride) with appropriate amines in benzene.
B) Construction of amide bond followed by ester linkage based on
retrosynthetic route b.2-10
Esterification of carboxylic acid which includes NSAIDs with
appropriate N-substituted 2-chloroacetamide in DMF at 25 °C 2,7 or 90 °C
3-8, 10 in presence of sodium iodide and TEA or in presence of catalytic
amount of DMAP9 using TEA as base.
3.3 OBJECTIVE OF THE PRESENT WORK
N,N-Disubstituted glycolamide esters of NSAIDs1 are extensively
used as carrier linked prodrug as they are hydrolysed extremely rapidly
in human plasma solutions, however, the rate of plasma catalysed
hydrolysis can be altered with the change of substituents on amide
nitrogen atom. Monosubstituted (-CO2CH2CONHR) or unsubstituted (-
CO2CH2CONH2) glycolamide esters were found to be more resistant than
N,N-disubstituted glycolamide esters (-CO2CH2CONRR').
Moreover, the glycolamide esters of aspirin failed to deliver aspirin in
human plasma when morpholine was the corresponding amine of amide
moiety.5
Attracted by these ideas, we became interested in the synthesis and
pharmacological evaluation of library of compounds. Our intention was
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111
to identify small molecule as cytotoxic agent having attractive chemical
scaffold for the development of new cytotoxic agent.
COX-2 inhibitor nimesulide shows anticancer effects in several
cancer cells and due to our interest in chemical modification of
nimesulide,14 we felt that nimesulide based substituted glycolamide ester
moiety might be worth exploring as it is capable of forming additional
hydrogen bonds.
We designed new glycolamide analogues 14 by using nimesulide (13)
as starting scaffold (figure 3.2).
Figure 3.2: Design of new nimesulide based glycolamide esters 14
The following few pages describe our efforts in this direction and
delineate successful synthesis of the nimesulide based glycolamide
esters.
3.4 RESULTS AND DISCUSSION
It was initially planned to synthesize target compounds 14 by a two
step process as shown in scheme 3.1.
N-(4-Amino-2-phenoxyphenyl)methane sulfonamide 15 was
prepared15 by reducing 13. Chloroacetylation of 15 in CHCl3 in presence
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112
of TEA gave the desired intermediate 2-chloro-N-(4-
methanesulfonylamino-3-phenoxy-phenyl)-acetamide (16) in good yields.
Scheme 3.1: Synthesis of nimesulide based glycolamide esters 14
The chloro compound 16 was well characterized by using mass, IR,
1H NMR, and 13C NMR spectroscopic techniques. Mass spectrum (figure
3.3) showed a protonated molecular ion peak at m/z 355. Appearance of
isotopic peak at m/z 357 confirmed the presence of one chlorine atom.
Figure 3.3: Mass (+Ve) spectrum of 16
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113
IR spectrum (figure 3.4) displayed a strong band at 1665 cm-1 due to
C=O stretching of amide group. The NH stretching frequency appeared at
ν 3325 and 3265 cm–1.
Figure 3.4: IR spectrum of 16
1H NMR spectrum in DMSO-d6 (figure 3.5) displayed two singlets at δ
4.20 and 2.96 ppm due to CH2 and CH3 groups respectively. When D2O
exchange experiment was performed two signals at δ 10.46 and 9.22
ppm disappeared which was in consistent with the presence of two labile
hydrogens (NH groups) in the structure.
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114
Figure 3.5: 1H NMR (DMSO-d6, 400 MHz) spectrum of 16
Figure 3.6: 13C NMR (CDCl3, 100 MHz) spectrum of 16
Appearance of total 13 singlets in 13C NMR spectrum (BB mode) of
compound 16 (figure 3.6) confirmed the presence of thirteen chemically
non equivalent carbon atoms. Peak at δ 164.7 ppm confirmed the amide
carbon. The signal for methyl group appeared at δ 39.4 ppm. The signal
at δ 52.7 ppm was due to CH2 group.
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115
Based on these spectral data and synthetic sequence the proposed
structure of 16 was confirmed. Detailed spectral data for 16 has been
given in the experimental section 3.7.1 of this chapter (page no. 126).
The second step, which is reaction of the resulting compound 16 with
commercially available acids (17a-l), was performed in the presence of
catalytic amount of DMAP9 using TEA as a base. To our surprise instead
of the desired glycolamide esters we isolated a white solid 18 with
melting point 264-265 °C irrespective of acid partner. All spectral data
which includes mass, IR, 1H NMR & 13C NMR were recorded and was
confirmed as 4-(dimethylamino)-1-(2-oxo-2-(4-methanesulfonylamino-3-
phenoxy phenylamino) ethyl) pyridinium salt (18) (scheme 3.2).
NHSO2Me
OPh
HN
O
Cl
NHSO2Me
OPh
HN
O
O R
O
NHSO2Me
OPh
HN
O
N
NMe2
+
RCOOH
DMAP, TEA rt
16 18
14
17
Scheme 3.2: Formation of pyridinium salt 18
Mass spectrum displayed molecular ion peak appeared at m/z 441
(figure 3.7).
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116
Figure 3.7: Mass spectrum of 18
Figure 3.8: IR spectrum of 18
IR spectrum of 18 (figure 3.8) displayed absorption at 1652 cm–1 due
to C=O stretching of amide group. In 1H NMR spectrum (figure 3.9),
appearance of singlet at δ 3.19 ppm, equivalent to 6H was indicative.
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117
Figure 3.9: 1H NMR (DMSO-d6, 400 MHz) spectrum of 18
Figure 3.10: 13C NMR (DMSO-d6, 100 MHz) spectrum of 18
In 13C NMR (figure 3.10) total seventeen signals appeared for
seventeen chemically non equivalent carbon atom including δ 164.7 for
amide carbon and δ 40.4 for geminal carbon atom of two methyl groups.
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118
Due to insolubility of the compound further nucleophilic attack by
carboxylate ion could not take place.
Hence we decided to synthesize the glycolamide esters in a different
way.1 As shown in scheme 3.3 nimesulide based glycolamide esters 14
were synthesized by one pot sequential nucleophilic substitution of the
key intermediate 16 to 2-iodo-N-(4-methanesulfonylamino-3-phenoxy-
phenyl)-acetamide in the presence of potassium iodide and finally to the
glycolamide esters with appropriate and commercially available acids 17
in the presence of base TEA.
Scheme 3.3: Synthesis of 14
Acetic acid (17a) was chosen as acid partner to establish the
optimized condition. Then the acid (0.01 mol) was reacted with the key
intermediate 16 (0.01 mol) in presence of potassium iodide (0.001mol),
TEA (0.011 mol) and 10 mL DMF at 90 °C. The product (acetic acid (4-
methanesulfonylamino-3-phenoxy-phenyl carbamoyl)-methyl ester) (14a)
isolated was characterized by mass, IR and NMR spectroscopic analysis.
In mass spectrum (figure 3.11) protonated molecular ion peak
appeared at m/z 379 corresponding to the molecular formula
C17H18N2O6S.
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119
Figure 3.11: Mass (+Ve) spectrum of 14a
The presence of two carbonyl groups in the product was confirmed by
its IR spectrum which showed carbonyl stretching frequencies at ν 1742
cm–1 for ester and 1677 cm–1 for amide (figure 3.12).
Figure 3.12: IR spectrum of 14a
In 1H NMR spectrum (figure 3.13) the appearance of two singlets at δ
2.20 ppm for 3H & δ 4.64 for 2H confirmed the presence of –COMe group
and CH2 group respectively.
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120
Figure 3.13: 1H NMR (CDCl3, 400 MHz) spectrum of 14a
In 13C NMR (figure 3.14) appearance of 15 signals were highly
consistent with the 15 non equivalent carbon atoms of the product. Ester
and amide carbonyls appeared at δ 169.9 & 165.5 ppm respectively,
where as CH2 appeared at δ 62.4 ppm.
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121
Figure 3.14: 13C NMR (DMSO-d6, 100 MHz) spectrum of 14a
Having established the optimum reaction conditions, we then
decided to examine the reaction of 16 with other carboxylic acids 17b-l
including NSAIDs. All glycolamide esters 14a-l synthesized were isolated
in good to excellent yields and formations of no side products were
detected. The results are summarized in table 3.1.
Table 3.1 Comparison of time and yield of products 14a-l.
Entry Ar/R-COOH Ar/R= (17)
Products (14) Time (min)
Yield (%)
1
Me 17a
NHSO2Me
OPh
HN
O
O Me
O
14a
30 91
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122
2
Et 17b
NHSO2Me
OPh
HN
O
O Et
O
14b
30 95
3 Ph 17c
14c
20 80
4
17d
14d
30 90
5
17e
14e
30 70
6
17f
14f
15 75
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123
7
17g
14g
10
98
8
17h
14h
30
78
9
17i
14i
45 80
10
HN
Cl
Cl 17j
NHSO2Me
OPh
HN
O
O
O
HN
Cl
Cl 14j
15 92
11
17k
14k
30 90
12
17l
14l
15 90
Chapter-3
124
3.5 CYTOTOXIC ACTIVITY
The in vitro cytotoxic evaluation of the tested glycolamide ester
derivatives against human HCT-15 for colon cancer compared to that of
doxyrubicin as the reference drug is shown in table 3.2. It should be
noted that compound 14l exhibited the highest toxicity.
Table 3.2 Cytotoxic activity of synthesized compounds 14a-l
S.No Compounds % of cell death at various concentrations
1 µg
/mL
2 µg
/mL
5 µg
/mL
10 µg
/mL
25 µg
/mL
1 14a 0.94 6.89 8.15 8.77 20.30
2 14b 0.62 7.20 7.50 18.50 38.50
3 14c 0.31 12.5 2.50 7.52 31.00
4 14d 3.43 4.53 21.69 35.78 49.38
5 14e 1.88 6.58 13.16 17.55 28.80
6 14f 5.88 14.46 20.22 25.00 42.40
7 14g 6.00 7.96 14.46 18.62 41.91
8 14h 0.31 7.52 8.77 12.20 19.40
9 14i 7.35 12.37 14.95 16.29 48.89
10 14j 6.74 9.43 25.61 35.90 51.22
11 14k 0 5.96 12.20 30.70 55.79
12 14l 21.69 21.93 24.50 30.02 66.05
All the values are the average of the experiments done in triplicates. The cell line used was HCT-15 human colon cancer cell line. Doxorubicin [IC50 = 50µg /mL (0.09 µM)] was used as reference compound. IC50 value of 14k and l are 21.5 and 18.4 respectively (figure 3.15).
Chapter-3
125
14k 14l Figure 3.15: In vitro cytotoxic activity of few synthesized compounds
against HCT-15, human colon cancer cell line.
3.6 CONCLUSION
In conclusion, we have successfully accomplished the synthesis of
several new nimesulide based glycolamide esters in good yields.
Structures of the synthesized compounds were confirmed by
spectroscopic analysis. The new derivatives were examined in vitro for
their cytotoxic activities. Our results indicate that the compounds
possess low to moderate cytotoxic activity against HCT-15 human colon
cancer cell line.
3.7 EXPERIMENTAL SECTION
All the carboxylic acids used are commercially available.
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126
3.7.1 Synthesis of 2-chloro-N-(4-methanesulfonylamino-3-phenoxy-
phenyl)-acetamide (16)
NHSO2Me
OPh
HN
O
Cl
1 g (3.6 mmol) of compound 15 was taken into a round bottom flask.
To this, 20 mL of CHCl3 and 0.6 mL (4.3 mmol) of TEA were added. This
mixture was stirred and then cooled to 0 oC. To this mixture 0.3 mL (3.6
mmol) of X-chloroacetyl chloride was added drop wise. The mixture was
then allowed to come to room temperature and stirring was continued for
an additional 30 minutes. After completion of the reaction as monitored
by TLC, the reaction mixture was quenched with 10-15 mL of cold water,
which was then extracted with CHCl3. The organic layers were collected,
combined, washed with water, dried over anhydrous Na2SO4 and
concentrated under reduced pressure. The crude was recrystallised with
aqueous EtOH.
DESCRIPTION: Off white solid.
M.P: 135-136 °C.
RF: 0.45 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3325, 3265, 1665, 1611, 1589.
Chapter-3
127
1H NMR (400 MHz, DMSO-d6): δ 10.30 (s, 1H, NH, D2O exchangeable),
9.25 (s, 1H, NH, D2O exchangeable), 7.50-7.40 (m, 2H), 7.38-7.26 (m,
2H), 7.21-7.14 (m, 2H), 7.10 (d, J ═ 8.8 Hz, 2H), 4.20 (s, 2H), 3.00 (s,
3H).
13C NMR (100 MHz, CDCl3): δ 164.7, 155.4, 148.2, 134.8, 130.2, 124.7,
124.4, 123.1, 118.8, 115.6, 110.1, 52.7, 39.4.
MASS (m/z): 355 [M+H]+, (47.5 %).
ELEMENTAL ANALYSIS found C, 50.84; H, 4.02; N, 7.73.
C15H15ClN2O4S requires C, 50.78; H, 4.26; N, 7.90 %.
3.7.2 General procedure for synthesis of glycolamide esters, 14a-l
A mixture of N-chloroacetamide (16) (3.54 g, 10 mmol), appropriate
acid (17) (10 mmol), KI (0.166 g, 1 mmol) and TEA (1.53 mL, 11 mmol) in
10 mL DMF was stirred at 90 °C. The progress of the reaction was
monitored by TLC. After completion, the reaction mixture was poured
into water (50 mL) and extracted with ethyl acetate (3×50 mL). The
combined organic extracts were washed with aqueous sodium
bicarbonate (2 %, 50 mL) and water (3×50 mL). The organic layer was
dried over anhydrous sodium sulphate and evaporated under reduced
pressure to get solid residue which was purified by column
chromatography to get the corresponding glycolamide esters 14a-l.
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128
3.7.2.1 Acetic acid (4-methanesulfonylamino-3-phenoxy-phenyl
carbamoyl)-methyl ester (14a)
NHSO2Me
OPh
HN
O
O Me
O
DESCRIPTION: Light orange solid.
M.P: 116-120 °C.
RF: 0.38 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3277, 1742, 1677, 1616, 1591.
1H NMR (400 MHz, CDCl3): δ 7.78 (bs, 1H, NH, D2O exchangeable), 7.58
(d, J ═ 8.8 Hz, 1H), 7.41-7.37 (m, 2H), 7.32 (d, J ═ 2.1 Hz, 1H), 7.21-7.13
(m, 2H), 7.01 (d, J ═ 7.9 Hz, 2H), 6.76 (s, 1H), 4.62 (s, 2H), 2.98 (s, 3H),
2.20 (s, 3H).
13C NMR (100 MHz, DMSO-d6): δ 169.9, 165.5, 155.8, 151.1, 137.8,
130.0, 127.8, 123.9, 122.9, 119.3, 114.0, 108.8, 62.4, 40.3, 20.4.
MASS (m/z): 379 [M+H]+, (100 %).
ELEMENTAL ANALYSIS found C, 53.84; H, 4.82; N, 7.33. C17H18N2O6S
requires C, 53.96; H, 4.79; N, 7.40 %.
Chapter-3
129
3.7.2.2 Propionic acid (4-methanesulfonylamino-3-phenoxy-phenyl
carbamoyl)-methyl ester (14 b)
NHSO2Me
OPh
HN
O
O Et
O
DESCRIPTION: Light orange solid.
M.P: 102-104 °C.
RF: 0.52 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3278, 2928, 1747, 1674, 1612.
1H NMR (400 MHz, CDCl3): δ 7.72 (bs, 1H, NH, D2O exchangeable), 7.59
(d, J ═ 8.5 Hz, 1H), 7.41-7.37 (m, 2H), 7.34 (s, 1H), 7.19-7.17 (m, 1H),
7.13 (dd, J ═ 8.5, 1.8 Hz, 1H), 7.01 (d, J ═ 8.3 Hz, 2H), 6.78 (bs, 1H, NH,
D2O exchangeable), 4.63 (s, 2H), 2.98 (s, 3H), 2.48 (q, J ═ 7.6 Hz, 2H),
1.20 (t, J ═ 7.6 Hz, 3H).
13C NMR (100 MHz, DMSO-d6): δ 173.2, 165.5, 155.9, 151.0, 137.4,
130.0, 127.8, 123.9, 122.9, 119.2, 114.0, 108.8, 62.3, 40.3, 26.4, 8.8.
MASS (m/z): 393 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 55.39; H, 5.02; N, 7.01. C18H20N2O6S
requires C, 55.09; H, 5.14; N, 7.14 %.
Chapter-3
130
3.7.2.3 Benzoic acid (4-methanesulfonylamino-3-phenoxy-phenyl
carbamoyl)-methyl ester (14c)
NHSO2Me
OPh
HN
O
O Ph
O
DESCRIPTION: White solid.
M.P: 127-128 °C.
RF: 0.72 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3266, 2933, 1736, 1672, 1611, 1548.
1H NMR (400 MHz, CDCl3): δ 8.07 (d, J ═ 7.4 Hz, 2H), 7.77 (s, 1H), 7.65-
7.61 (m, 2H), 7.59-7.50 (m, 2H), 7.48-7.36 (m, 3H), 7.20-7.16 (m, 1H),
7.10 (dd, J ═ 8.7, 2.2 Hz, 1H), 7.01 (d, J ═ 8.0 Hz, 2H), 6.73 (s, 1H), 4.88
(s, 2H), 2.97 (s, 3H).
13C NMR (100 MHz, DMSO-d6): δ 165.4, 155.8, 151.1, 137.4, 133.6,
130.0, 129.3, 129.1, 128.8, 127.9, 123.9, 122.9, 119.3, 114.0, 108.7,
63.0, 40.3.
MASS (m/z): 441 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 60.09; H, 4.34; N, 6.17. C22H20N2O6S
requires C, 59.99; H, 4.58; N, 6.36 %.
Chapter-3
131
3.7.2.4 2-Methyl-benzoic acid (4-methanesulfonylamino-3-phenoxy
phenylcarbamoyl)-methyl ester (14d)
NHSO2Me
OPh
HN
O
O
O Me
DESCRIPTION: White solid.
M.P: 139-140 °C.
RF: 0.51 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3274, 3236, 1727, 1677, 1613, 1224, 1153.
1H NMR (400 MHz, CDCl3): δ 7.95 (dd, J ═ 8.7, 1.5 Hz, 1H), 7.78 (s, 1H),
7.60 (d, J ═ 8.2 Hz, 1H), 7.48 (dd, J ═ 7.6, 1.5 Hz, 1H), 7.41 -7.37 (m,
3H), 7.35-7.29 (m, 2H), 7.17 (t, J ═ 8.1 Hz, 1H), 7.10 (dd, J ═ 8.7, 2.0 Hz,
1H), 7.02 (d, J ═ 7.7 Hz, 2H), 6.72 (s, 1H), 4.86 (s, 2H), 2.97 (s, 3H), 2.68
(s, 3H).
13C NMR (100 MHz, CDCl3): δ 180.0, 165.7, 165.3, 155.5, 148.1, 140.9,
134.7, 133.0, 132.1, 131.5, 130.2, 127.9, 126.0, 124.6, 123.0, 118.6,
115.8, 110.6, 63.3, 39.4, 21.8.
MASS (m/z): 455 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 60.56; H, 4.69; N, 6.27. C23H22N2O6S
requires C, 60.78; H, 4.88; N, 6.16 %.
Chapter-3
132
3.7.2.5 4-Methoxy-benzoic acid (4-methanesulfonylamino-3-
phenoxy phenylcarbamoyl)-methyl ester (14e)
NHSO2Me
OPh
HN
O
O
O
OMe
DESCRIPTION: White solid.
M.P: 198-200 °C.
RF: 0.64 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3340, 3307, 2843, 1710, 1604.
1H NMR (400 MHz, CDCl3): δ 8.03 (d, J ═ 8.8 Hz, 2H), 7.76 (bs, 1H, NH,
D2O exchangeable), 7.60 (d, J ═ 8.4 Hz, 1H), 7.39 (t, J ═ 8.5 Hz, 3H),
7.20-7.17 (m, 1H) 7.09 (dd, J ═ 8.8, 2.2 Hz, 1H), 7.01 (d, J ═ 8.6 Hz, 2H),
6.97 (d, J ═ 8.8 Hz, 2H), 6.71 (bs, 1H, NH, D2O exchangeable), 4.86 (s,
2H), 3.89 (s, 3H), 2.97 (s, 3H).
13C NMR (100 MHz, DMSO-d6): δ 165.6, 165.0, 163.3, 155.8, 151.1,
137.4, 131.5, 130.0, 127.9, 123.9, 122.9, 121.3, 119.3, 114.0, 108.8,
62.7, 55.5, 40.3.
MASS (m/z): 471 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 58.58; H, 4.60; N, 6.12. C23H22N2O7S
requires C, 58.71; H, 4.71; N, 5.95 %.
Chapter-3
133
3.7.2.6 4-Nitro-benzoic acid (4-methanesulfonylamino-3-phenoxy-
phenylcarbamoyl)-methyl ester (14f)
NHSO2Me
OPh
HN
O
O
O
NO2
DESCRIPTION: White solid.
M.P: 206-208 °C.
RF: 0.49 (CHCl3 : EtOAc = 7 : 3).
IR (KBr) νmax/cm–1: 3273, 3096, 2949, 1728, 1682, 1663.
1H NMR (400 MHz, DMSO-d6): δ 10.33 (s, 1H, NH, D2O exchangeable),
9.26 (s, 1H, NH, D2O exchangeable), 8.37 (d, J ═ 8.8 Hz, 2H), 8.22 (d, J ═
8.7 Hz, 2H), 7.43 (t, J ═ 7.8 Hz, 2H), 7.36-7.27 (m, 2H), 7.22 (d, J ═ 3.0
Hz, 1H), 7.18 (t, J ═ 7.3 Hz, 1H), 7.07 (d, J ═ 7.3 Hz, 2H), 4.92 (s, 2H),
3.32 (s, 3H).
13C NMR (100 MHz, CDCl3 + DMSO-d6): δ 163.9, 163.0, 155.3, 149.5,
148.9, 136.0, 133.8, 130.0, 128.9, 125.3, 122.8, 122.7, 122.5, 117.8,
114.0, 109.1, 62.6, 39.1.
MASS (m/z): 486 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 54.54; H, 4.23; N, 8.78. C22H19N3O8S
requires C, 54.43; H, 3.94; N, 8.66 %.
Chapter-3
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3.7.2.7 3-Phenyl-acrylic acid (4-methanesulfonylamino-3-phenoxy-
phenylcarbamoyl)-methyl ester (14g)
NHSO2Me
OPh
HN
O
O
O
Ph
DESCRIPTION: Buff solid.
M.P: 198-200 °C
RF: 0.62 (CHCl3 : EtOAc = 8 : 2).
IR (KBr) νmax/cm–1: 3251, 3092, 1719, 1682, 1509.
1H NMR (400 MHz, DMSO-d6): δ 10.22 (s, 1H, NH, D2O exchangeable),
9.25 (s, 1H, NH, D2O exchangeable), 7.75-7.68 (m, 3H), 7.45-7.40 (m,
4H), 7.35-7.28 (m, 3H), 7.23 (d, J ═ 2.4 Hz, 1H), 7.18 (t, J ═ 7.3 Hz, 1H),
7.07 (d, J ═ 7.4 Hz, 2H), 6.72 (d, J ═ 16.1 Hz, 1H), 4.72 (s, 2H), 2.96 (s,
3H).
13C NMR (100 MHz, DMSO-d6 + CDCl3): δ 174.3, 165.0, 164.7, 155.3,
148.8, 144.8, 136.1, 133.1, 129.6, 129.0, 128.0, 127.12 125.2, 122.9,
122.7, 117.9, 116.2, 114.2, 109.3, 61.8, 39.7.
MASS (m/z): 467 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 61.86; H, 4.59; N, 5.91. C24H22N2O6S
requires C, 61.79; H, 4.75; N, 6.00 %.
Chapter-3
135
3.7.2.8 2-(6-Methoxy-naphthalen-2-yl)-propionic acid (4-methane
sulfonylamino-3-phenoxy-phenylcarbamoyl)-methyl ester
(14h)
NHSO2Me
OPh
HN
O
O
O
Me
OMe
DESCRIPTION: Off white solid.
M.P: 196-198 °C
RF: 0.84 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3464, 3386, 3060, 2936, 1742, 1692, 1607.
1H NMR (400 MHz, CDCl3): δ 7.70 (d, J ═ 9.1 Hz, 2H), 7.67 (bs, 1H, NH,
D2O exchangeable), 7.41-7.33 (m, 4H), 7.21-7.12 (m, 4H), 6.91 (d, J ═
8.0 Hz, 2H), 6.82 (s, 1H), 6.58 (s, 1H), 5.81 (dd, J ═ 8.8, 2.2 Hz, 1H),
4.87 (d, J ═ 15.8 Hz, 1H), 4.38 (d, J ═ 15.7 Hz, 1H), 3.98 (q, J ═ 7.0 Hz,
1H), 3.96 (s, 3H), 2.89 (s, 3H), 1.65 (d, J ═ 7.3 Hz, 3H).
13C NMR (50 MHz, DMSO-d6): δ 184.6, 173.5, 164.2, 162.5, 158.3,
157.1, 146.4, 135.4, 133.2, 129.2, 128.3, 126.8, 126.3, 125.7, 121.2,
118.6, 116.6, 115.8, 112.4, 105.7, 102.9, 62.7, 55.1, 52.2, 44.1, 18.5.
MASS (m/z): 549 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 63.62; H, 5.02; N, 4.98. C29H28N2O7S
requires C, 63.49; H, 5.14; N, 5.11 %.
Chapter-3
136
3.7.2.9 2-(4-Isobutyl-phenyl)-propionic acid (4-methanesulfonyl
amino-3-phenoxy-phenylcarbamoyl)-methyl ester (14i)
NHSO2Me
OPh
HN
O
O
O
Me
Me
Me
DESCRIPTION: Off white solid.
M.P: 48-50 °C.
RF: 0.50 (CHCl3).
IR (KBr) νmax/cm–1: 3336, 3284, 2954, 2931, 2868, 1746, 1693, 1612,
1590, 1537.
1H NMR (400 MHz, CDCl3): δ 7.50 (d, J ═ 8.7 Hz, 1H), 7.38 (t, J ═ 7.7
Hz, 2H), 7.26-7.07 (m, 7H), 6.97 (d, J ═ 7.6 Hz, 2H), 6.66 (s, 1H), 6.60
(d, J ═ 7.7 Hz, 1H), 4.82 (d, J ═ 15.7 Hz, 1H), 4.41 (d, J ═ 15.7 Hz, 1H),
3.81 (q, J ═ 7.0 Hz, 1H), 2.93 (s, 3H), 2.42 (d, J ═ 6.9 Hz, 2H), 1.85-1.79
(m, 1H),1.54 (d, J ═ 7.3 Hz, 3H), 0.89 (d, J ═ 6.2 Hz, 6H).
13C NMR (50 MHz, CDCl3): δ 172.5, 165.2, 155.6, 147.6, 141.5, 137.1,
134.6, 130.2, 129.9, 127.1, 124.5, 122.9, 118.2, 115.6, 110.7, 62.4,
44.9, 44.8, 39.3, 30.2, 22.3, 17.7.
MASS (m/z): 525 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 64.23; H, 6.02; N, 5.60. C28H32N2O6S
requires C, 64.10; H, 6.15; N, 5.34 %.
Chapter-3
137
3.7.2.10 [2-(2, 6-Dichloro-phenylamino)-phenyl]-acetic acid (4-
methanesulfonylamino-3-phenoxy-phenylcarbamoyl)-
methyl ester (14j)
NHSO2Me
OPh
HN
O
O
O
HN
Cl
Cl
DESCRIPTION: Off white solid.
M.P: 122-123 °C.
RF: 0.57 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3475, 3382, 3246, 1718, 1666, 1228, 1160.
1H NMR (400 MHz, CDCl3): δ 7.51 (d, J ═ 8.7 Hz, 1H), 7.38-7.33
(m, 4H), 7.28-7.14 (m, 4H), 7.06-6.87 (m, 6H), 6.69 (s, 1H, NH, D2O
exchangeable), 6.42 (d, J ═ 8.2 Hz, 1H), 6.28 (s, 1H, NH, D2O
exchangeable), 4.69 (s, 2H), 3.92 (s, 2H), 2.95 (s, 3H).
13C NMR (100 MHz, CDCl3): δ 180.1, 169.7, 164.7, 155.7, 147.7, 142.4,
137.0, 134.5, 130.8, 130.2, 129.9, 128.9, 128.7, 124.9, 124.5, 122.9,
122.7, 122.2, 118.5, 117.9, 115.7, 110.3, 62.9, 39.4, 38.2.
MASS (m/z): 614 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 56.34; H, 4.01; N, 6.52.
C29H25Cl2N3O6S requires C, 56.68; H, 4.10; N, 6.84 %.
Chapter-3
138
3.7.2.11 2-(2, 3-Dimethyl-phenylamino)-benzoic acid (4-
methanesulfonyl amino-3-phenoxy-phenylcarbamoyl)-
methyl ester (14k)
NHSO2Me
OPh
HN
O
O
O HN
Me
Me
DESCRIPTION: Pale yellow solid.
M.P: 140-141 °C.
RF: 0.50 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3338, 2925, 1687, 1608, 1508, 1327, 1218, 1156,
1098.
1H NMR (400 MHz, CDCl3): δ 9.08 (1H, NH), 7.96 (dd, J ═ 8.0, 1.1 Hz,
1H), 7.80 (bs, 1H, NH, D2O exchangeable), 7.61 (d, J ═ 8.4 Hz, 1H), 7.41-
7.05 (m, 9H), 7.01 (d, J ═ 7.7 Hz, 2H), 6.75-6.68 (m, 3H), 4.88 (s, 2H),
2.97 (s, 3H), 2.33 (s, 3H), 2.15 (s, 3H).
13C NMR (50 MHz, DMSO-d6): δ 167.2, 165.5, 155.7, 151.0, 148.6,
138.0, 137.9, 137.3, 134.7, 131.6, 131.5, 129.9, 127.8, 126.7, 126.0,
123.9, 122.9, 122.7, 119.2, 116.3, 113.9, 113.3, 109.9, 108.8, 62.7,
40.7, 20.2, 13.5.
MASS (m/z): 560 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 64.15; H, 5.09; N, 7.60. C30H29N3O6S
requires C, 64.39; H, 5.22; N, 7.51 %.
Chapter-3
139
3.7.2.12 2-Acetoxy-benzoic acid (4-methanesulfonylamino-3-
phenoxy-phenylcarbamoyl)-methyl ester (14l)
NHSO2Me
OPh
HN
O
O
O O Me
O
DESCRIPTION: Off white solid.
M.P: 147-148 °C.
RF: 0.39 (CHCl3 : EtOAc = 9 : 1).
IR (KBr) νmax/cm–1: 3249, 3154, 3099, 2944, 1758, 1730, 1674, 1610,
1555, 1507.
1H NMR (400 MHz, DMSO-d6): δ 10.28 (s, 1H, NH, D2O exchangeable),
9.26 (s, 1H, NH, D2O exchangeable), 8.00 (d, J ═ 6.5 Hz, 1H), 7.71 (t, J ═
6.5 Hz, 1H), 7.45-7.16 (m, 8H), 7.07 (d, J ═ 8.0 Hz, 2H), 4.81 (s, 2H),
2.96 (s, 3H), 2.24 (s, 3H).
13C NMR (100 MHz, DMSO-d6): δ 169.1, 165.1, 163.5, 155.8, 152.9,
151.1, 150.2, 137.3, 134.4, 130.1, 127.9, 126.3, 124.2, 124.0, 122.9,
122.4, 119.3, 113.9, 108.7, 63.1, 40.3, 20.7.
MASS (m/z): 499 [M+H]+, (100 %).
ELEMENTAL ANALYSIS: found C, 57.69; H, 4.49; N, 5.38. C24H22N2O8S
requires C, 57.82; H, 4.45; N, 5.62 %.
Chapter-3
140
3.7.2.13 4-(Dimethylamino)-1-(2-oxo-2-(3-phenoxy phenylamino)
ethyl) pyridinium (18) (scheme 3.2)
NHSO2Me
OPh
HN
O
N
NMe
Me
+
DESCRIPTION: White solid.
M.P: 264-265 °C.
IR (KBr) νmax/cm-1: 3408, 3175, 3045, 1652, 1607, 1332, 1221.
1H NMR (400 MHz, DMSO-d6): δ 10.93 (s, 1H, NH, D2O exchangeable),
9.30 (bs, 1H, NH, D2O exchangeable), 8.20 (d, J ═ 7.6 Hz, 2H), 7.32-7.29
(m, 3H), 7.17-6.98 (m, 5H), 6.85 (d, J ═ 7.9 Hz, 2H), 5.13 (s, 2H), 3.19 (s,
6H ), 2.96 (s, 3H).
13C NMR (100 MHz, DMSO-d6): δ 164.7, 155.9, 155.8, 151.1, 143.3,
137.4, 130.0, 127.8, 124.0, 123.1, 119.2, 114.0, 108.9, 107.1, 58.3,
40.4, 38.9.
MASS (m/z): 441 (M+, 100 %).
3.8 REFERENCES
1. Bundgaard, H.; Nielsen, N. M. J. Med. Chem., 30, 1987, 451–454.
2. Nielsen, N. M.; Bundgaard, H. J. Pharm. Sci., 77, 1988, 285–298.
3. Wadhwa, L. K.; Sharma, P. D. Ind. J. Chem., 34B, 1995, 408–415.
Chapter-3
141
4. Sharma, P. D.; Singh, K. J.; Gupta, S.; Chandiran, S. Indian. J.
Chem., 43B, 2004, 636–642.
5. Nielsen, N. M.; Bundgaard, H. J. Med. Chem., 32, 1989, 727–734.
6. Bansal, A. K.; Khar, R. K.; Dubey, R.; Sharma, A. K. Drug Dev. Ind.
Pharm., 27, 2001, 63–70.
7. Gadad, A. K.; Bhat, S.; Tegli, V. S.; Redasani, V. V. Arzneimittel-
Forschung, 52, 2002, 817–821.
8. Cao, F.; Gueo, J. X.; Ping, Q. N.; Liao, Z. G. Eur. J. Pharm., 29,
2006, 385–393.
9. Smriti, K.; Manjula, M.; Akhila, V.; Rahul, B.; Ram, T.; Basha, S.
K. J. S.; Ramesh, M.; Rao, C. S.; Pal, M. Bioorg. Med. Chem., 14,
2006, 4820–4833.
10. Nalini, C. N.; Ramachandran, S.; Kavitha, K.; Saraswathi, V. S. Int.
J. Res. Pharm. Biomed. Sci., 2, 2011, 1112–1117.
11. Amblard, M.; Rodriguez, M.; Martinez, J. Tetrahedron, 44, 1988,
5101–5108.
12. Hoekstra, W. J. US 6066651.
13. Boltze, K. H.; Kreisfeld, H. Arzneimittel-Forschung, 27, 1977, 1300–
1312.
14. (a) Kavitha, K.; Reddy, V. R.; Mukkanti, K.; Pal, S. J. Braz. Chem.
Soc., 21, 2010, 1060–1064. (b) Abir, B.; Ghosh, S.; Kavitha, K.;
Reddy, V. R.; Mukkanti, K.; Pal, S.; Mukherjee, A. K. Chem. Phy.
Lett., 493, 2010, 151–157. (c) Kavitha, K.; Varsha, P.; Mukkanti,
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K.; Reddy, V. R.; Pal, S. Molbank, 2011, M740; doi:
10.3390/M740.
15. Pericherla, S.; Mareddy, J.; Rani, D. P. G.; Gollapudi, P. V.; Pal, S.
J. Braz. Chem. Soc., 18, 2007, 384–390.
3.9 SOME IMPORTANT SPECTRA OF THE COMPOUNDS
Figure 3.16: Mass (+Ve) spectrum of 14b
Figure 3.17: IR spectrum of 14b
Chapter-3
143
Figure 3.18: 1H NMR (CDCl3, 400 MHz) spectrum of 14b
Figure 3.19: 13C NMR (DMSO-d6, 100 MHz) spectrum of 14b
Chapter-3
144
Figure 3.20: 1H NMR (CDCl3, 400 MHz) spectrum of 14c
Figure 3.21: 13C NMR (DMSO-d6, 100 MHz) spectrum of 14c
Chapter-3
145
Figure 3.22: 1H NMR (CDCl3, 400 MHz) spectrum of 14d
Figure 3.23: 13C NMR (CDCl3, 100 MHz) spectrum of 14d
Chapter-3
146
Figure 3.24: IR spectrum of 14e
Figure 3.25: 1H NMR (CDCl3, 400 MHz) spectrum of 14e
Chapter-3
147
Figure 3.26: 1H NMR (DMSO-d6, 400 MHz) spectrum of 14f
Figure 3.27: 13C NMR (CDCl3 + DMSO-d6, 100 MHz) spectrum of 14f
Chapter-3
148
Figure 3.28: 1H NMR (DMSO-d6, 400 MHz) spectrum of 14g
Figure 3.29: 13C NMR (CDCl3 + DMSO-d6, 100 MHz) spectrum of 14g
Chapter-3
149
Figure 3.30: 1H NMR (CDCl3, 400 MHz) spectrum of 14h
Figure 3.31: 13C NMR (DMSO-d6, 50 MHz) spectrum of 14h
Chapter-3
150
Figure 3.32: 1H NMR (CDCl3, 400 MHz) spectrum of 14i
Figure 3.33: 13C NMR (CDCl3, 50 MHz) spectrum of 14i
Chapter-3
151
Figure 3.34: 1H NMR (CDCl3, 400 MHz) spectrum of 14j
Figure 3.35: 13C NMR (CDCl3, 100 MHz) spectrum of 14j
Chapter-3
152
Figure 3.36: 1H NMR (CDCl3, 400 MHz) spectrum of 14k
Figure 3.37: 13C NMR (DMSO-d6, 50 MHz) spectrum of 14k
Chapter-3
153
Figure 3.38: 1H NMR (DMSO-d6, 400 MHz) spectrum of 14l
Figure 3.39: 13C NMR (DMSO-d6, 100 MHz) spectrum of 14l