Development of amino acid conjugated sulfonamides as potent antiulcer agent

Article abstract of DOI:10.1007/s00044-013-0891-y

A series of 2-{[(4-methylphenyl) sulfonyl] amino}-3-sulfanylpropanoic acid (1a) and its analogs 1b-j have been synthesized. These compounds were screened for their in vivo efficacy in pyloric ligation model. Compounds 1a and 1b with higher antiulcer potential were further screened in other gastric models to explore the mode of antiulcer action. To further understand the mode of action, in vitro inhibition of H⁺/K⁺ ATPase activity in gastric microsome isolated from rat stomach was studied. This was rationalized by in silico experiments.

Full text of DOI:10.1007/s00044-013-0891-y

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-013-0891-y
ORIGINAL RESEARCH
Development of amino acid conjugated sulfonamides as potent
antiulcer agent
•
Shakti Prasanna Sahoo Bharat Bhusan Subudhi
Received: 26 August 2013 / Accepted: 11 December 2013
Ó Springer Science+Business Media New York 2013
Abstract A series of 2-{[(4-methylphenyl) sulfonyl]
amino}-3-sulfanylpropanoic acid (1a) and its analogs
1b–j have been synthesized. These compounds were
screened for their in vivo efficacy in pyloric ligation model.
Compounds 1a and 1b with higher antiulcer potential were
further screened in other gastric models to explore the
mode of antiulcer action. To further understand the mode
of action, in vitro inhibition of H?/K? ATPase activity in
gastric microsome isolated from rat stomach was studied.
This was rationalized by in silico experiments.
efficacious and more selective drugs to treat gastric ulcer is
vital in its chemotherapy.
Gastric carbonic anhydrase (CA) catalyzes the reaction
between carbon dioxide and water to form carbonic acid
which is immediately dissociated to release hydrogen ions
leading to transportation across parietal cell by H?/K?
ATPase located in the cannalicular membrane. Experi-
mental evidence has led researchers to suggest that H?/K?
ATPase and CA in the gastric mucosa are functionally
coupled and drugs like omeprazole has a dual mechanism
of action including H?/K? ATPase and gastric mucosa CA
inhibition (Puscas et al., 1999a, b, 2000; Purkerson and
Schwartz, 2005). Sulfonamides are well known for their
capacity to inhibit CA. In our previous investigations, we
have found sulfonamide to potentiate the antiulcer drug
action of 1,4-dihydropyridine and quinazoline derivatives
(Subudhi et al., 2009, 2012). However, the mechanism of
sulfonamides beyond CA inhibition is not clear, and there
is not enough work to establish their other modes of action.
In our group, we have been trying to derivatise amino
acids for developing potent antiulcer agents (Subudhi and
Sahoo 2011a, b). Amino acids including L-glutamic acid,
Glycine, L-tryptophan, L-Arginine, and L-serine have been
demonstrated to inhibit gastric secretion and protect the
gastric mucosa against chemical and stress-induced ulcers
(Tariq and Moutaery, 1997; Kalia et al., 2000; Ohta and
Nishida, 2002). Derivatives of amino acids including
dimethylglycine have been shown to have potent anti-
ulcer properties (Hariganesh and Prathiba, 2000). Several
N-acyl derivatives of a-amino acids were also found to be
effective inhibitors of gastric proton pump (Sharma et al.,
2007). Considering the potential shown by amino acids in
reducing ulceration, we developed amino acid derived
sulfonamides in our attempt to develop potent antiulcer
agents.
Keywords Amino acid ꢀ Sulfonamide ꢀ Conjugate ꢀ
Antiulcer ꢀ H?/K? ATPase
Introduction
Gastric ulcer is a chronic inflammation of the stomach
which may lead to gastric cancer (Marshall and Windsor,
2005).The existing antiulcer agents control ulceration but
provoke adverse effect on long term use. Besides owing to
a variety of reasons, the number of new gastric ulcer
patient continues to rise. Therefore, the quest for more
S. P. Sahoo
GIET School of Pharmacy, Chaitanya Knowledge City,
Rajahmundry 533296, Andhra Pradesh, India
e-mail: sahoo.shakti@gmail.com
B. B. Subudhi (&)
School of Pharmaceutical Sciences, Siksha O Anusandhan
University, Bhubaneswar 751003, Orissa, India
e-mail: bharatbhusans@gmail.com
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Med Chem Res
R
O
S
H2N
H3C
Cl
O
O
NaOH(aq)
HO
O
S
R
H3C
NH
O
O
1a-j
HO
Comp
R
Comp
R
Comp
R
1a
1b
1c
S
SH
N
H
1d
1e
1f
1i
NH
N
OH
CH3
O
1g
1j
1h
OH
CH3
NH2
OH
CH3
Scheme 1 Synthesis of amino acid conjugates of sulfonamide (1a–j)
Results and discussion
the reported method with little modification (Boer and
Backer, 1954). The starting material p-toluenesulfonyl
chloride was added to aqueous solution of amino acids
under alkaline condition. The pKa value of a-amino groups
of amino acids used in this experiment varies in a range of
8.8 for asparagine to 10.25 for cysteine. Accordingly,
during the reaction, care was taken to maintain the reaction
Chemistry
We prepared 2-{[(4-methylphenyl) sulfonyl] amino}-3-
sulfanylpropanoic acid (1a) and its analog (1b–j) according
to Scheme 1. Compounds were synthesized according to
123

Med Chem Res
mixture at a pH around 11–12 and in excess of it, in order
to maintain an uncharged a-amino group with enhanced
nucleophilicity. The test compounds (1a–j) were recovered
and purified following appropriate conditions by column
chromatography. The yield was highest for compound 1a
(75 %) and lowest for 1e (39 %). The fewer yield in case of
1e can be ascribed to the formation of sulfonate as
byproduct because of the hydroxyl group of tyrosine. All
compounds were confirmed by physicochemical properties,
1a, 1b, 1g, and 1h have similar ability to reduce acid
secretion only in case of 1a and 1b, there was higher
neutralization of acid as evident from a better pH. Besides
these compounds exhibited better effectiveness as reflected
in lower values of free and total acidity as well as lower
scores of ulcer index. Based on these findings and histo-
pathology of mucosal layer of animals (Fig. 1) treated with
1a and 1b, these compounds were selected for further
investigation in different models to explore the spectrum of
their activity. In order to generate more evidence, the
gastroprotective effect was evaluated in aspirin, alcohol
(AL), and cold-resistance stress-induced ulcer model.
Besides the ability of test compounds to inhibit gastric, H?/
K? ATPase was tested and rationalized by in silico
experiments.
1
FTIR, H NMR, 13C NMR, Mass, and elemental analysis.
The FTIR spectrum of 1b exhibited a broad peak at
3,446 cm-1 (O–H Str.) suggesting carboxylic group. This
1
was supported by C=O stretching at 1,693 cm-1. The H
NMR spectrum of the compound 1b consists of charac-
teristic singlet for methyl substitution on aromatic ring and
NH proton at d 2.35 and d 8.033, respectively. Character-
istic multiplet for aromatic protons was observed in the
range of d 7.17–7.67. This was further supported by 13C
NMR spectrum exhibiting peaks at d 176.8 and d 59.1
assigned to carbon of carbonyl (acid) and methanetriyl
group, respectively. Besides molecular ion peak at 303(m/
e) in mass spectrum of 1b followed by elemental analysis
confirmed the proposed structure. Specific rotations of
compounds suggested their optical properties because of
the chiral centers present in the compounds.
Aspirin-induced ulceration
This type of ulceration is attributed principally to inhibition
of biosynthesis of cytoprotective prostaglandins, e.g.,
PGE’s and PGI2 (by inhibition of cyclooxygenase pathway
of arachidonic acid metabolism), resulting in overproduc-
tion of leukotrienes and other products of 5-lipoxygenase
pathway (Rainsford, 1987). Leukotriene antagonist and
5-lipoxygenase inhibitors are capable of inhibiting NSA-
IDs-induced gastric ulceration in rats (Parnaham and
Brune, 1987). Thus, protection of test compounds 1a and
1b (Table 2) in this model against ulceration may indicate
their ability to inhibit 5-lipoxygenase pathway or to possess
leukotriene antagonist activity. Reduction in gastric vol-
ume and the acid neutralizing activity for 1a and 1b in this
model were found significant (P \ 0.001). Compound 1a
has shown more decrease in ulcer index as well as free and
total acidity compared to 1b (Table 2).
Pharmacology
Oral acute toxicity study of the synthesized compounds
was done by guidelines of Organization of Economic Co-
operation and Development (OECD-423) for testing of
chemicals. Lethality was not observed till 2,000 mg/kg
indicating the safety of the compounds (1a–j). Antiulcer
activity was studied in pyloric ligation model with ome-
prazole as standard drug (10 mg/kg) and reported in
Table 1 (Shay et al., 1945; Ishii et al., 1976; Nair et al.,
2010). Reduction in volume of gastric juice was found to
be comparable to that of omeprazole for the compounds 1a,
1b, 1g, and 1h. The acid neutralizing ability as indicated by
pH of the gastric fluid was found to be more for compounds
1a, 1b, and 1c. All the compounds exhibited significant
(P \ 0.001) acid neutralizing capacity. Compound 1a, 1b,
1g, and 1h showed good antisecretory activity comparable
to omeprazole. Reduction in free acidity by all synthesized
compounds indicated their ability to inhibit secretion of
hydrochloric acid. All the compounds were observed to
inhibit free acidity and total acidity. The potential of the
test compounds for inhibiting hydrochloric acid, gastric
juice, and neutralizing ability has resulted in protection of
gastric mucosa as observed from the significant
(P \ 0.001) decrease in ulcer index and histopathological
studies (Fig. 1). It was observed that although compounds
Alcohol-induced gastric ulceration
Ethanol promotes serious disturbances in the gastric
mucosa leading to intense gastric ulcer (Guth et al., 1984;
Hiruma-Lima et al., 2009). It acts by exerting a direct toxic
effect on the epithelium, leading to the formation of
characteristic necrotic lesions due to a reduction in the
mucus production and bicarbonate secretion (Massignani
et al., 2009). In addition, it induces reduction of gastric
blood flow, the solubilization of components of the mucus
of the stomach, and oxidative stress, increase of xanthine
oxidase activity and levels of malondialdehyde with
reduction in levels of glutathione (Marrotta et al., 1999).
Thus, test compounds 1a and 1b displayed antiulcer effect
related to cytoprotective activity, since they significantly
reduced the ethanol-induced ulcer (Table 2).
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Med Chem Res
Table 1 Antiulcer activity of test compounds 1a–j by pyloric ligation model
Group
Volume (ml)
pH
Ulcer index
Free acidity (mEq./l)
Total acidity (mEq./l)
Control
10.6 0.49
2.38 0.24a
4.82 0.46a
4.78 0.46a
6.05 0.31a
4.43 0.56a
5.38 0.44a
6.42 0.73a
4.28 0.53a
4.68 0.47a
6.25 0.64a
5.02 0.55a
1.86 0.29
5.47 0.52a
5.03 0.53a
5.62 0.51a
5.25 0.40a
4.15 0.35a
4.03 0.43a
4.67 0.39a
4.22 0.39a
4.25 0.36a
4.27 0.62a
4.18 0.58a
21.3 1.41
2.98 0.52a
8.18 1.09a
9.77 0.78a
9.60 1.18a
9.05 1.17a
10.6 0.51a
10.5 0.59a
12.4 1.37a
11.8 0.66a
9.36 0.61a
9.13 1.45a
57.67 2.34
9.100 0.65a
15.00 1.84a
17.67 1.66a
20.67 2.04a
22.50 2.43a
24.67 2.86a
25.50 2.07a
21.00 3.05a
24.83 2.48a
30.83 3.83a
24.50 2.26a
62.82 3.07
10.50 0.77a
24.00 3.13a
23.67 3.04a
27.00 2.47a
25.83 2.53a
29.50 3.01a
30.33 2.49a
29.83 3.11a
28.67 3.50a
34.67 3.86a
27.83 2.12a
Omeprazole
1a
1b
1c
1d
1e
1f
1g
1h
1i
1j
Data are represented as mean SEM. Statistical analysis was done by one-way ANOVA followed by Dunnett’s Multiple Comparison Test
a
P \ 0.001 as compared to control (n = 6 in each group)
have been suggested to be the principal factor in stress-
Compound
Histopathology of gastric mucosal layer
induced ulceration and can be prevented partially or
entirely by vagotomy (Brodie and Hanson, 1960; Suleyman
et al., 2001). Accordingly, the protective action of 1a and
1b against stress-induced ulceration suggests a related
antiulcer effect (Table 2).
Control
In vitro assay of H?/K? ATPase activity
Standard
Gastric microsome isolated from rat stomach was analyzed
after incubation with test compounds and omeprazole
(Sanui, 1974; Berglindh, 1990). It was observed (Fig. 2)
that 1a and 1b inhibited the proton pump in a dose
dependent manner. About 78 % inhibition was shown by
1b at a dose of 50 lg/ml and 72 % inhibition by 1a which
are significant as comparing to the standard drug (91 %).
These observations were in agreement with our previous
findings in different ulcer model. It is thus possible that the
strong gastric acid secretion inhibitory properties observed
in vivo for 1a and 1b is due to intricate mechanisms as
discussed above correlating their acid neutralizing capacity
with gastro-protective, H?/K? ATPase inhibitory as well
as anti-secretory properties within the stomach.
1a
1b
In silico studies
Fig. 1 Histopathology of gastric mucosal layer (pyloric ligation
model)
The druglikeness study does not show any violations as per
the Lipinski’s rule (Table 3). This suggests their likliness
to be potential drug candidate (Lipinski, 2004). The
potential of 1a and 1b against ulceration was rationalized
by in silico experiments with auto dock vina wizard (Trott
and Olson, 2010). The chiral center present in the test
compounds and stereo specificity of drug target interaction
R and S configuration of test compounds were considered
Cold restraint stress-induced gastric ulceration
In this model, gastric ulcer formation is mainly due to
gastric hyper motility associated with enhanced acid
secretion and a reduction in mucous production (Qiu et al.,
1992). Vagal over-activity and excess histamine release
123

Med Chem Res
Table 2 Antiulcer activity of 1a and 1b in different ulcer models
Group
Volume (ml)
pH
Ulcer index
Free acidity
Total acidity
Aspirin-induced model
Control
10.3 0.54
3.28 0.26a
5.76 0.42a
5.58 0.50a
2.34 0.24
20.9 1.01
3.01 0.12a
8.15 1.01a
9.76 0.78a
53.67 2.34
10.2 0.95a
15.83 1.77a
17.83 1.93a
59.43 2.835
12.5 0.47a
24.17 3.27a
25.33 2.98a
Omeprazole
6.30 0.32a
5.2 0.43a
5.65 0.48a
1a
1b
Alcohol-induced model
Control
Omeprazole
1a
11.1 0.61
2.85 0.14a
6.05 0.33a
4.63 0.53a
2.32 0.22
5.49 0.58a
5.4 0.33a
4.36 0.31a
21.0 1.33
3.47 0.82a
9.41 1.19a
7.88 0.89a
57.67 2.34
11.2 0.45a
22.5 1.92a
22.5 2.17a
63.82 2.42
12.7 0.87a
28.67 1.49a
30.83 1.95a
1b
Cold restraint stress-induced model
Control
Omeprazole
1a
9.02 0.52
2.89 0.44a
5.76 0.42a
5.85 0.55a
2.25 0.22
6.02 0.72a
4.61 0.29a
4.833 0.51a
20.2 1.36
3.21 0.45a
9.81 0.77a
9.45 0.47a
52.03 3.12
7.89 0.87a
24.67 2.2a
27.67 2.75a
58.82 3.12
10.21 0.32a
32.83 2.98a
36.0 4.12a
1b
Data are represented as mean SEM. Statistical analysis was done by one-way ANOVA followed by Dunnett’s multiple comparison test
a
P \ 0.001 as compared to control (n = 6 in each group)
including cysteine, methionine, and p-methyl benzene
sulfonamide exhibited low affinity for the target suggesting
the fact their conjugation as in case of test compounds has
been able to enhance their interaction with H?/K? ATPase.
This is in agreement with the experimental findings.
Experimental
Chemistry
All reagents and solvents were procured and used without
further purification. Melting points were determined on a
Fig. 2 H?/K? ATPase inhibition activity 1a (filled triangle), 1b (filled
circle), omeprazole (filled square)
Sisco melting point apparatus and are uncorrected. Crude
products were purified by column chromatography on silica
gel of 60–120 mesh. Molecular weights of the compounds
for this study. Compound 1a and 1b showed affinity
comparable to that of Omeprazole (Table 4). Both R and S
configuration of test compounds exhibited close binding
affinity. The ligand-target interaction visualized for R
configuration of both ligand shows very close interaction
(Fig. 3). The integral fragments of test compounds
were determined by Rast’s procedure. IR spectra were
obtained on a JASCO FTIR-4100 spectrometer using KBr
pellet. NMR spectra were recorded on BRUKER
AVANCE-II-400 MHz spectrometer using DMSO. The
chemical shifts were reported as ppm down field using
tetramethylsilane (TMS) as an internal standard. Elemental
Table 3 Drug likeness data of 1a and 1b
2
TPSA (A )
3
Volume (A )
˚
˚
Compound
LogP
No. of atoms
MW (g)
ON
OHNH
No. of violations
Rotatable bonds
1a
1b
0.448
0.027
83.468
83.468
17.0
19.0
275.351
303.405
5
5
2
2
0
0
5
7
222.73
256.56
Log P Partition coefficient, TPSA Molecular polar surface area (square angstroems), MW Molecular weight (g), ON No of hydrogen bond
acceptors, OHNH No of hydrogen bond donors, Volume Molecular volume(cubic angstroems)
123

Med Chem Res
analysis was carried out with PERKIN ELMER-2400
analyzer. Mass spectra were recorded on a MICROMASS
Q-TOF MICRO spectrometer. Specific rotations of com-
pounds were measured by observation of their angle of
rotation using LAB-276 polarimeter.
cysteine (174 ml of the 40 % solution) in a round-bot-
tomed flask. The mixture was allowed to heat up to
80–90 °C in order to maintain the sulfonyl methyl amide in
a molten condition. The mixture was shaken vigorously.
Boiling was prevented by mild cooling with water in order
to avoid an excessive loss of methylamine. As soon as the
reaction mixture became acidic as indicated by testing a
drop on litmus paper, 50 ml of the 50 % sodium hydroxide
solution was added carefully with swirling. If the mixture
became acidic again 25 ml of 50 % sodium hydroxide
solution is added to make the final mixture alkaline. The
walls of the flask were rinsed with a little water. The
reaction was completed by heating the mixture consisting
of two layers and a precipitate of sodium chloride on a
steam bath for 15 min with vigorous mechanical stirring.
The hot reaction mixture was then poured into glacial
acetic acid contained in a round-bottomed flask. Then the
solution was cooled in an ice bath to about 5 °C and stirred
mechanically. To this, a solution of sodium nitrite (1.8 mol
in 250 ml of water) was added from a dropping funnel
during about 45 min. The temperature of the mixture was
kept below 10 °C, and stirring was continued for 15 min
after addition was complete. During the reaction, the final
compound separated as a crystalline product. The crystal-
line product was washed with 500 ml of water. This was
further recrystallised from methanol followed by purifica-
tion by column chromatography using methanol and ben-
zene (2:1) as mobile phase. The final dried compounds
were kept in desiccator for further analysis. Other
Procedure for 2(R)-{[(4-methylphenyl) sulfonyl]
amino}-3-sulfanylpropanoic acid (1a)
p-Toluene sulfonyl chloride (1.68 mol) was added with
swirling during about 5 min to 2.1 mol of 33 % aqueous
Table 4 In-silico binding affinity of test compounds against H?/K?
ATPase (2XZB)
Compounds
Binding affinity (Kcal/mol)
1a(R)
-6.5
-6.3
-6.5
-6.5
-6.9
-4.4
-4.8
-5.7
1a(S)
1b(R)
1b(S)
Omeprazole
Cysteine
Methionine
PMBS
Binding affinity in kcal/mol with root mean square deviation of upper
bound and lower bound as zero.2XZB is the X-ray crystallographic
structure of Pig gastric H?/K? ATPase.R & S represent the ligand
configuration. PMBS is p-methyl benzene sulfonamide
Fig. 3 Molecular docking of 1a
and 1b with gastric H?/K?
ATPase (2XZB)
Comp
Surface representation
Polar interaction
1a
1b
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Med Chem Res
compounds of the series (1b–j) were obtained in similar
manner.
N, 13.58; found: C, 50.17; H, 4.59; N, 13.38. IR (KBr),
v (cm-1): 3431 (O–H), 3334 (N–H), 2991 (Ar–H), 2749
(C–H), 1682 (C=O), 1355 (S–H). 1H NMR (DMSO) d
(ppm): 11.23 (brs,1H,OH), 11.02(d,1H,NH) 7.34–7.32(m,
6H,Ar–H), 4.25(d,1H,NH), 3.70(m,1H,CH), 3.17(d,2H,
CH2). 13C NMR (CDCl3) d (ppm):136.2 (C2-imidazole),
131.3 (C4-imidazole), 117.2 (C5-imidazole), 27.5 (C6-
methanediyl), 56.4 (C7-methanetriyl), 174.4 (C11-car-
bonyl), 141.3 (C14–Ar), 128.3 (C15–Ar), 129.3(C16–Ar),
137.6 (C17–Ar), 21.2 (C21–methyl). MS (m/z): 309 (M?).
2(S)-{[(4-methylphenyl)sulfonyl]amino}-3-
sulfanylpropanoic acid (1a)
Brown solid.Yield: 75 %. Mp: 124–126 °C. Analysis for
C10H17NO4S2 (275.344); calculated: C, 43.62; H, 4.76; N,
5.09; found: C, 43.22; H, 4.36; N, 5.01. IR (KBr), v (cm-1):
3420 (O–H), 3350 (N–H), 2991 (Ar–H), 2754 (C–H), 1692
25
1
(C=O),1367 (S–H). H NMR (DMSO) d (ppm): 7.16–7.73
½aꢁD : -46.
(m,4H,Ar–H), 3.67–3.67–3.86 (m,2H,CH2), 2.34 (s,3H,CH3).
13C NMR (CDCl3) d (ppm): 141 (C1–Ar), 128.2 (C2–Ar),
129.5 (C3–Ar), 139.3 (C6–Ar), 21.4 (C9-methyl), 58.9 (C12-
methanetriyl), 25.1 (C13-methanediyl), 177.2 (C15-carbonyl).
3-(4-Hydroxyphenyl)-2(S)-{[(4-methylphenyl)sulfonyl]
amino}propanoic acid (1e)
25
MS m/z (%): 276 (M?). ½aꢁD : -18.
Brown solid. Yield: 39 %: mp 130–132 °C. Analysis for
C16H17NO5S (335.374); calculated: C, 57.30; H, 5.11; N,
4.18; found: C, 57.27; H, 5.06; N, 4.12. IR (KBr), v (cm-1):
3426 (O–H), 3356 (N–H), 2981 (Ar–H), 2774 (C–H), 1692
(C=O), 1362 (S–H). 1H NMR (DMSO) d (ppm): 11.27
(brs,1H,OH), 7.34–7.30 (m,8H,Ar–H),5.7 (s,1H,OH), 4.22
(d,1H,NH), 3.80 (m,1H,CH), 2.78 (d,2H,CH2), 2.34
(s,3H,CH3). 13C NMR (CDCl3) d (ppm): 141.2 (C1–Ar),
128.3 (C2–Ar), 129.2(C3–Ar), 137.6 (C4–Ar), 21.3 (C9-
methyl), 57.2 (C12-methanetriyl), 35.4 (C13-methanediyl),
130.3 (C15–Ar), 115.6 (C16–Ar), 155.5 (C17–Ar), 174.7
2(S)-{[(4-methylphenyl)sulfonyl]amino}-4-
(methylsulfanyl)butanoic acid (1b)
White solid. Yield: 64 %. Mp: 133–135 °C. Analysis for
C12H17NO4S2 (303.397); calculated: C, 47.50; H, 5.65; N, 4.62;
found: C, 47.43; H, 5.35; N, 4.42. IR (KBr), v (cm-1): 3446 (O–
H), 3350 (N–H), 2981 (Ar–H), 2754 (C–H), 1693 (C=O), 1358
(S–H). 1H NMR (DMSO) d (ppm): 7.17–7.67 (m,4H,Ar–H),
3.9 (d,1H,NH), 2.59–2.61 (m,1H,CH), 2.35 (s,3H,CH3), 2.07
(s,3H,CH3). 13C NMR (CDCl3) d (ppm): 141.4 (C1–Ar), 128.2
(C2–Ar), 129.3 (C3–Ar), 139.2 (C6–Ar), 21.4 (C9-methyl), 59.1
(C12-methanetriyl), 28.5 (C13-methanediyl), 29.2.5 (C14-
methanediyl), 16.2 (C16-methyl), 176.8 (C17-carbonyl). MS (m/
25
(C20-carbonyl). MS (m/z): 335 (M?). ½aꢁD : -52.
2(S)-{[(4-methylphenyl)sulfonyl]amino}-3-
phenylpropanoic acid (1f)
25
z): 303 (M?). ½aꢁD : ?126.
White solid.Yield 56 %. Mp: 141–143 °C. Analysis for
C16H17NO4S (319.375); calculated: C, 60.17; H, 5.37; N,
4.39; found: C, 60.12; H, 5.27; N, 4.29. IR (KBr), v (cm-1):
3426 (O–H), 3341(N–H), 2989 (Ar–H), 2754 (C–H), 1690
(C=O), 1370 (S–H). 1H NMR (DMSO) d (ppm): 11.26
(brs,1H,OH), 7.31–7.28(m,9H,Ar–H), 4.20–4.18(d,1H,NH),
3.80(m,1H,CH), 2.90–2.88 (d,2H,CH2), 2.32(s,3H,CH3).
13C NMR (CDCl3) d (ppm): 141.2 (C1–Ar), 128.2 (C2–Ar),
129.2 (C3–Ar), 137.4 (C4–Ar), 21.3 (C9-methyl), 57.4 (C12-
methanetriyl), 35.6 (C13-methanediyl), 136.3 (C14–Ar),
127.1 (C15-Ar), 128.7 (C16-Ar), 125.4 (C17–Ar), 174.7 (C20-
3-(1H-indol-3-yl)-2(S)-{[(4-methylphenyl)sulfonyl]
amino}propanoic acid (1c)
Off white solid. Yield: 72 %. Mp: 136–138 °C. Analysis for
C18H18N2O4S (358.411); calculated: C, 60.32; H, 5.06; N, 7.82;
found: C, 60.12; H, 4.96; N, 7.62. IR (KBr), v (cm-1): 3427 (O–
H), 3340 (N–H), 2998 (Ar–H), 2751 (C–H), 1678 (C=O), 1360
1
(S–H). H NMR (DMSO) d (ppm): 11.18 (s,1H,OH), 9.76
(d,1H,NH), 7.17–7.12 (m,9H,Ar–H), 4.22 (d,1H,NH), 3.80
(m,1H,CH), 2.76 (d,2H,CH2), 2.30 (s,3H,CH3). 13C NMR
(CDCl3) d (ppm):122.7 (C2-indole), 109.5 (C3-Indole), 127.2
(C4-indole), 118.4 (C5-indole), 119.2 (C6-indole), 121.4 (C7-
indole),111.2 (C8-indole), 136.3 (C9-indole), 29.1 (C10-meth-
anediyl), 58.3 (C11-methanetriyl), 174.4 (C15-carbonyl), 141.2
(C18–Ar), 128.3 (C19–Ar), 129.3 (C20–Ar), 137.2 (C21–Ar),
25
carbonyl). MS (m/z): 319 (M?). ½aꢁD : -44.
3-Hydroxy-2(S)-{[(4-methylphenyl)sulfonyl]amino}
propanoic acid (1g)
25
21.3 (C25-methyl). MS (m/z): 358 (M?). ½aꢁD : -68.
Off white solid. Yield 53 %. Mp: 127–129 °C. Analysis for
C10H13NO5S (259.278); calculated: C, 46.32; H, 5.05; N,
5.40; found: C, 46.22; H, 4.95; N, 5.36. IR (KBr), v (cm-1):
3417 (O–H), 3350 (N–H), 29877 (Ar–H), 2761 (C–H),
1602 (C=O), 1368 (S–H). 1H NMR (DMSO) d (ppm):
11.22 (s,1H,OH), 7.43–7.40 (d,4H,Ar–H), 4.18 (d,1H,NH),
3-(1H-imidazol-4-yl)-2(S)-{[(4-methylphenyl)sulfonyl]
amino}propanoic acid (1d)
Paleyellow solid. Yield: 55 %. Mp: 117–119 °C. Analysis
for C13H15N3O4S (309.34); calculated: C, 50.47; H, 4.89;
123

Med Chem Res
4.02 (t,1H,OH), 3.58 (m,1H,CH), 3.50(t,2H,CH2), 2.54(s,
3H,CH3). 13C NMR (CDCl3) d (ppm): 141.3 (C1–Ar),
128.2 (C2–Ar), 129.2 (C3–Ar), 137.7 (C4–Ar), 21.3 (C9-
methyl), 58.1 (C12-methanetriyl), 59.4 (C13-methanediyl),
174.6 (C15-carbonyl), 19.2 (C17-methyl). MS (m/z): 273
25
(M?). ½aꢁD : -15.
Pharmacology
25
174.3 (C15-carbonyl). MS (m/z): 259 (M?). ½aꢁD : ?148.
All the synthesized compounds were evaluated for antiul-
cer activity using white albino rats following the reported
method (Shay et al., 1945; Ishii et al., 1976; Nair et al.,
2010) using omeprazole (1 mg/kg) as standard. Prior to
induction of ulcer, the animals were fasted with free access
to water for a period of 24 h so as to ensure complete
gastric emptying and a steady state gastric acid secretion.
The fasted animals were administered with the drugs/
vehicle on the morning of the experiment. After 1 h, the
animals were anaesthetized using anesthetic ether, and a
midline incision was made just below the xiphoid process.
The stomach was lifted out and ligated at the level of the
pylorus following which it was replaced and the abdomen
wall was closed by interrupted sutures. The animals were
kept deprived of food and water and were sacrificed by an
overdose of anesthetic ether after 4 h. At the end of
experiment, the animals were sacrificed by euthanasia, and
the stomach was removed. Gastric contents were drained
from the stomach and centrifuged at 3,000 rpm for 5 min,
and the volume of supernatant solution was measured. Free
and total acidity were estimated by titration with 0.1 N
NaOH using topfer reagent and phenolphthalein as indi-
cator. Acid output was expressed in mEq by multiplying
the volume in ml by the acid concentration in mEq/l. For
estimation of ulcer index inner surface of the stomach was
examined for ulceration with magnifying glass. The dif-
ference between the groups was determined using the one-
way analysis of variance (ANOVA).
3-Methyl-2(S)-{[(4-methylphenyl)sulfonyl]amino}
butanoic acid (1h)
White solid. Yield: 44 %. Mp: 141–143 °C. Analysis for
C12H17NO4S (271.332); calculated : C, 53.12; H, 6.32; N,
5.16; found: C, 53.02; H, 6.22; N, 5.06. Found: C, 53.02;
H, 6.22; N, 5.06. IR (KBr), v (cm-1): 3441 (O–H),
3362(N–H), 2951 (Ar–H), 2774 (C–H), 1662 (C=O), 1358
(S–H). 1H NMR (DMSO) d (ppm): 11.22 (brs,1H,OH),
7.43–7.40(m,4H,Ar–H), 4.21(d,1H,NH), 3.42(m,1H,CH),
2.37–2.35(m,1H,CH), 2.30(s,3H,CH3), 1.56(s,6H,CH3).
13C NMR (CDCl3) d (ppm): 141.2 (C1–Ar), 128.2 (C2–Ar),
129.3 (C3–Ar), 137.6 (C4–Ar), 21.3 (C9-methyl), 61.7 (C12-
methanetriyl), 29.3 (C13-methanetriyl), 18.7 (C14-methyl),
25
174.5 (C15-carbonyl). MS (m/z): 271 (M?). ½aꢁD : ?110.
5-Amino-2(S)-{[(4-methylphenyl)sulfonyl]amino}-5-
oxopentanoic acid (1i)
White solid.Yield: 58 %. Mp: 131–133 °C. Analysis for
C12H16N2O5S(300.33);calculated:C,47.99;H, 5.37;N,9.33;
found: C, 47.79; H, 5.34; N, 9.13. IR (KBr), v (cm-1): 3451
(O–H), 3337(N–H), 29967 (Ar–H), 2764 (C–H), 1692 (C=O),
1372 (S–H). 1H NMR (DMSO) d (ppm): 11.31(brs,1H,OH),
7.31–7.28(dd,4H,Ar–H), 4.24(d,1H,NH),4.16(s,2H,NH2)
3.22–3.19(d,4H,CH2), 2.34(s,3H,CH3). 13C NMR (CDCl3)
d (ppm): 141.3 (C1–Ar), 128.3 (C2–Ar), 129.2(C3–Ar), 137.2
(C4–Ar), 21.3 (C9-methyl), 54.1 (C12-methanetriyl), 25.3
(C13-methanediyl), 33.2 (C14-methanediyl), 173.5 (C15-car-
Aspirin-induced (ASP) gastric ulcer model in rats
25
bonyl), 174.7(C17-carbonyl). MS(m/z):300(M?).½aꢁD :-65.
Aspirin at a dose of 150 mg/kg, po was administered to
induce ulcer after 45 min of administration of 1a, 1b
(50 mg/kg), and omeprazole (10 mg/kg, po). The animals
were sacrificed 5 h after aspirin treatment (Goel et al.,
1985; Parnaham and Brune, 1987), and the stomach was
dissected out and the lesion was scored.
3(S)-Hydroxy-2(S)-{[(4-methylphenyl)sulfonyl]amino}
butanoic acid (1j)
Off white solid.Yield: 59 %. Mp: 131–137 °C. Analysis for
C11H15NO5S (273.305); calculated : C, 48.34; H, 5.53; N,
5.12; found: C, 48.14; H, 5.33; N, 5.02 IR (KBr), v (cm-1):
3455 (O–H), 3363(N–H), 2951 (Ar–H), 2744 (C–H), 1672
(C=O), 1363 (S–H). 1H NMR (DMSO) d (ppm): 11.29
(brs,1H,OH), 7.34–7.31 (m,4H,Ar–H), 4.22 (d,1H,NH),
4.12 (d,1H,OH) 3.62 (m,2H,CH), 2.32(s,3H,CH3), 1.92
(d,3H,CH3). 13C NMR (CDCl3) d (ppm): 141.5 (C1–Ar),
128.3 (C2–Ar), 129.3 (C3–Ar), 137.2 (C4–Ar), 21.3 (C9-
methyl), 62.4 (C12-methanetriyl), 65.3 (C13-methanetriyl),
Alcohol (AL) induced gastric ulcer model in rats
Gastric ulcer was induced in rats by administering absolute
AL (1 ml/200 g, body, 1 h). Compounds 1a, 1b, and
sucralfate (500 mg/kg, po) were administered 45 min
before AL treatment (Suleyman et al., 2002). After 1 h, the
animals were sacrificed, and stomachs were excised to
observe the gastric lesions and were measured.
123

Med Chem Res
Cold restraint stress (CRU)-induced gastric ulcer model
in rats
secretory as well as proton pump inhibitory properties that
contribute to their antiulcer action and make them suitable
leads for further optimisation.
The rats were subjected to cold-stress paradigm, after
45 min of treatment with 1a, 1b, and omeprazole (10 mg/
kg, po). All animals were immobilized in a restraint cages,
kept at 4 °C in an environmental chamber for 2 h and
sacrificed thereafter. The stomach was cut along the lesser
curvature, and ulcers were scored. The stomach was dis-
sected out, gastric contents were collected and subjected to
centrifugation (1,000 rpm for 10 min) and analyzed for
volume and pH (Suleyman et al., 2001).
Acknowledgments The authors are thankful to Director, sophisti-
cated analytical instrument facility, Punjab University, Chandigarh
for their kind help in analysis and Director, Pharmacology Division,
Central Drug Research Institute, Chattar Manzil Palace, Lucknow 226
001, India, For their kind help in evaluation of proton pump activity.
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