Synthesis, spectroscopic characterization, molecular docking and theoretical studies (DFT) of N-(4-aminophenylsulfonyl)-2-(4-isobutylphenyl) propanamide having potential enzyme inhibition applications

Article abstract of DOI:10.1016/j.bmc.2019.01.012

A mutual prodrug (1) of ibuprofen and sulphanilamide has been synthesized with dual activity and improved toxicity profile. The synthesized compound has been characterized by elemental analysis, FT-IR, ¹HNMR, ¹³CNMR and ESI-MS. The m

Full text of DOI:10.1016/j.bmc.2019.01.012

Accepted Manuscript
Synthesis, Spectroscopic Characterization, Molecular Docking and Theoretical
Studies (DFT) of N-(4-aminophenylsulfonyl)-2-(4-isobutylphenyl) propana-
mide having Potential Enzyme Inhibition Applications
Amina Asghar, Muhammad Yousuf, Hifsa Mubeen, Rabia Nazir, Kabiru
Haruna, Abdulmujeeb T. Onawole, Lubna Rasheed
PII:
S0968-0896(18)31649-3
https://doi.org/10.1016/j.bmc.2019.01.012
BMC 14704
DOI:
Reference:
To appear in:
Bioorganic & Medicinal Chemistry
Received Date:
Revised Date:
Accepted Date:
23 September 2018
17 December 2018
15 January 2019
Please cite this article as: Asghar, A., Yousuf, M., Mubeen, H., Nazir, R., Haruna, K., Onawole, A.T., Rasheed, L.,
Synthesis, Spectroscopic Characterization, Molecular Docking and Theoretical Studies (DFT) of N-(4-
aminophenylsulfonyl)-2-(4-isobutylphenyl) propanamide having Potential Enzyme Inhibition Applications,
Bioorganic & Medicinal Chemistry (2019), doi: https://doi.org/10.1016/j.bmc.2019.01.012
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Synthesis, Spectroscopic Characterization, Molecular Docking and Theoretical Studies (DFT)
of N-(4-aminophenylsulfonyl)-2-(4-isobutylphenyl) propanamide having Potential Enzyme
Inhibition Applications
a,
b
a
c
d,e
Amina Asghar , Muhammad Yousuf , Hifsa Mubeen , Rabia Nazir , Kabiru Haruna ,
d
a,*
Abdulmujeeb T. Onawole , Lubna Rasheed
a
Department of Chemistry, Division of Science and Technology, University of Education, Lahore,
Pakistan
Department of Chemistry, Ulsan National Institute of Science and Technology (UNIST), Ulsan,
b
South Korea.
c
Pakistan Council of Scientific and Industrial Research, Lahore, Pakistan.
Department of Chemistry, King Fahd University of Petroleum and Minerals, Dhahran 31261,
d
Saudi Arabia.
e
Department of Chemistry, Ahmadu Bello University, Zaria 1044, Nigeria.
Abstract
A mutual prodrug (1) of ibuprofen and sulphanilamide has been synthesized with dual
activity and improved toxicity profile. The synthesized compound has been characterized by
1
13
elemental analysis, FT-IR, HNMR, CNMR and ESI-MS. The molecular geometry of the
compound (1) was optimized using density functional theory (DFT/B3LYP) method with the 6-
3
11G(d, p) basis sets in ground state. Geometric parameters (bond lengths, bond angles, torsion
angles), vibrational assignments, chemical shifts and thermodynamics of the compound (1) has been
calculated theoretically and compared with the experimental data. Comparative AutoDock study of
compound (1) with cyclooxygenase enzymes (COX-1 and COX-2) were performed involving
docking for possible selectivity of our prodrug within the two Cox enzymes. The highest binding
affinities of -8.7 Kcal/mol and -8.1 Kcal/mol has been obtained for COX-1 and COX-2 enzymes
respectively. Compound (1) exhibited enhanced anti-inflammatory, anti-ulcer and free radical
scavenging activities as compared with the parent drugs. Based on various in vitro and in vivo tests it
is suggested that the Compound (1) is more active than the parent drugs. Moreover, LD of
5
0
compound (1) is higher than parent drug i.e. ibuprofen and sulphanilamide suggesting that the
synthesized compound is much safer than its parent analogous.
Keywords: Ibuprofen, Sulfanilamide, Antibiotics, Anti-inflammatory drugs, AutoDock, DFT
*
Corresponding author. Tel.: 92-307-0766743
E-mail addresses: lubnarasdheed@ue.edu.pk (L. Rasheed)

1
. Introduction
Treatment of fever and inflammation dates back to centuries (~ 400 B.C.) when the Greek
physician Hippocrates prescribed an extract from willow bark and leaves.[1] It was about 17th
century when the active ingredient of willow bark salicin was identified and acetylsalicyclic acid
(aspirin) was introduced into the market by Bayer in 1899. But the mechanism of action of anti-
inflammatory and analgesic agents such as aspirin was unclear until 1971 when John Vane
discovered the mechanism of action of aspirin and other nonsteroidal anti-inflammatory drugs
(NSAIDs) thereby increasing our ability to develop novel anti-inflammatory therapies.[2] It is well
known phenomenon that infections usually also result in inflammation. Therefore, it is common
clinical practice that non-steroidal anti-inflammatory drugs (NSAIDs) are concomitantly prescribed
with anti-infective.[3] Thus Non-steroidal anti-inflammatory drugs (NSAIDs) form a group of
compounds which serve the purpose of fixing/inhibiting cyclooxygenase and thereby arresting
prostaglandin production through arachidonic acid metabolism.[4]
Generally NSAIDs such as aspirin, ibuprofen and diclofenac exhibit nonselective COX inhibition
but these are most widely prescribed NSAIDs to relieve short term fever, pain and inflammation.[5]
The characteristic feature of these traditional nonselective COX inhibitor NSAIDs was the presence
of a carboxylic acid (COOH) functional group.[6] In the early 1990s the second isoform of COX was
discovered, providing a novel target to develop anti-inflammatory agents with superior safety
profiles compared to traditional NSAIDs.[7,8] Consequently, selective COX-2 inhibitors (coxibs)
based on a diarylheterocyclic ring template as in celecoxib and rofecoxib were developed.[9,10]
Scientists are taking interest in determination of the structural and spectroscopic properties of
compounds using both experimental techniques and theoretical methods to have better understanding
of phenomenon at molecular level. Moreover, autodock studies of targeted compound give us a clear
picture of interaction of a drug with enzyme of interest thus saving time that has been spent in past
for in vitro experiments.[11] Density functional theory (DFT) is a powerful tool in theoretical
modeling and with recent advancement in computational facilities it been able to have great accuracy
in reproducing the experimental values for the geometry, dipole moment, vibrational frequency,
etc.[12-19] Molecular docking is one of the common tools used in computer-aided drug designing
(CADD) which is widely used in studying the binding affinity of a ligand to a target protein and also
in understanding the possible mode of action by which the ligand inhibits the target protein in disease
treatment.[20] A comparison of the experimental and theoretical studies can be very useful in

making correct assignments and understanding the basic phenomenon and hence establishment of
molecular structure-activity relationship.
Pharmacologically and commercially important NSAIDs like aspirin, ibuprofen, naproxen,
indomethacin, flurbiprofen have been well studied both kinetically and structurally.[6-10] But
another distinct class of compounds, the sulphanilamide group, was not given proper attention for
time long. Keeping in view such a necessity, design and synthesis of mutual prodrugs (PD) involving
anti-infectives and NSAIDs has been of interest. This design carries the benefits of a prodrug, which
are: better lipophilicity, better bio-availability, reduced toxicity and a sustained release effect.
Keeping in view of above mentioned facts we have synthesized a novel prodrug of ibuprofen and
sulphanilamide, N-(4-aminophenylsulfonyl)-2-(4-isobutylphenyl) propanamide, (1). The synthesized
compound (1) has been characterized by instrumental techniques and molecular geometry was
optimized using density functional theory (DFT/B3LYP) method with the 6-311G(d,p) basis sets in
ground state. Geometric parameters (vibrational assignments, chemical shifts) has been calculated
theoretically and compared with the experimental data. Comparative AutoDock study of compound
(
1) was also conducted with cyclooxygenase enzymes (COX-1 and COX-2) involving docking for
possible selectivity of our prodrug within the two Cox enzymes. The highest binding affinities of -
.7 Kcal/mol and -8.1 Kcal/mol has been obtained for COX-1 and COX-2 enzymes respectively. The
8
compound was also subjected to in vitro analysis and the results showed enhanced anti-
inflammatory, anti-ulcer and free radical scavenging activities as compared with the parent drugs. To
the best of our knowledge, no experimental, computational and in vitro study on the compound (1)
has been published in the literature yet. Furthermore, the presented data can be helpful in context of
the further studies of prodrugs of ibuprofen and its derivatives.
2
. Results and Discussion
2
.1 Synthesis
Synthesis of the title compound (1) was carried out by converting ibuprofen to its acid
chloride by use of oxalyl chloride followed by coupling with sulphanilamide through an –NH group
2
as shown in Scheme 1. There are two possible products as a result of the coupling reaction. We have
isolated the compound (1) by column chromatography. (detail in supporting information)
2
2
.2 Computational analysis
.2.1 Quantum chemical calculations

All the calculations were performed by using Gaussian 09 package [21] and Gauss-View
molecular visualization software [22] on the personal computer without restricting any symmetry.
The structure was optimized by density functional theory (DFT)/B3LYP [23,24] method with 6-
3
11G(d,p) as basis sets. From the optimized geometry of the molecule (Figure 1), vibrational
assignments and chemical shifts of the title compound have been calculated theoretically and
compared with the experimental data. Besides, the frontier molecular orbital (FMOs), molecular
electrostatic potential (MEP), Mulliken population analysis and thermodynamic properties of the
compound were also investigated by theoretical calculation results.
2
.2.2 Vibrational spectra
Theoretical harmonic frequencies had been calculated using density functional theory
(DFT/B3LYP) method with the 6-311G(d,p) basis sets in ground state. Scaling factors of 0.982 [25]
was used for B3LYP/6-311G(d,p) levels. Theoretical FT-IR spectra of the compound (Figure 2)
showed that the results are in agreement with experimental. (supporting information)
2
.2.3 NMR spectra
1
13
The H and C NMR spectra of the compound (1) recorded using TMS as an internal
standard and chloroform (CDCl ) solvent. Chemical shifts of hydrogen and carbon atoms were
3
1
13
determined from the obtained spectra. GIAO H- and C- chemical shift calculations had been
carried out using the B3LYP methods with 6-311G(d,p) as basis set for the optimized geometries.
1
13
The H and C NMR chemical shifts were converted to the TMS scale by subtracting the calculated
absolute chemical shielding of TMS (δ = Σo - Σ), where δ is the chemical shift, Σ is the absolute
shielding and Σo is the absolute shielding of TMS, whose values are 31.64 and 193.08 ppm for
B3LYP/6-311G(d,p), respectively. To make comparison with experimental observations, we
presented correlation graphs in Figure 3 based on the calculations (correlation coefficients of 0.9994
1
13
for H NMR and 0.9984 for C NMR, respectively).
2
2
2.2.4 Quantum-chemical studies
.2.4.1 HOMO and LUMO analysis
Frontier molecular orbitals (FMO) i.e. highest occupied molecular orbital (HOMO) and
lowest lying unoccupied molecular orbital (LUMO) are important ones in a molecule. The HOMO
energy characterizes the ability of electron giving while LUMO energy characterizes the ability of
electron accepting. Molecular stability depends on the gap between HOMO and LUMO energy
levels [26]. The distributions of the HOMO and LUMO orbitals computed at the B3LYP/6-311G(d,
p) levels for the compound (1) shown in Figure 4. The calculations indicated that the compound (1)

has 76 occupied molecular orbitals and the value of the energy separation between the HOMO and
LUMO are -6.3085 and -1.1793 eV for at the same levels. By using HOMO and LUMO energy
values for a molecule, electronegativity, chemical hardness and chemical softness were calculated as
follows:
χ =(I+A)/2(electronegativity), η =(I-A)/2 (chemical hardness), S = 1/2 η (chemical softness). Where I
and A are ionization potential and electron affinity; I = -EHOMO and A = -ELUMO, respectively [27].
The HOMO and LUMO energies, the energy gap (ΔE), the ionization potential (I), the electron
affinity (A), the absolute electronegativity (χ), the absolute hardness (η) and softness (S) for molecule
had been calculated at the same levels and the results are given in Table 1.
2
.2.5 Molecular electrostatic potential (MEP)
A molecule of electrostatic potential map provides vital information about the electron
acceptor and electron-donor regions. Thus intramolecular and intermolecular hydrogen bonds can be
conveniently calculated. The different values of the electrostatic potential at the surface are
represented by different colors; red represents regions of most electro negative electrostatic potential,
blue represents regions of most positive electrostatic potential and green represents regions of zero
potential. The electrostatic potential increases in the order red < orange < yellow < green < blue.
Figure 5 showed the molecular electrostatic potential maps of compound for B3LYP/6-311G(d, p)
levels, where blue indicated the strongest attraction and red indicated the strongest repulsion. Here,
the red regions settled on O1, O2, and O3 atoms while blue at N2 atom respectively. So, it can be
estimated to be N2 atom electron-donor and O1, O2, and O3 atoms are electron-acceptor.
2
.2.6 Mulliken population analysis
The Mulliken charge distributions of the compound (1) was calculated using B3LYP/6-
11G(d, p) levels. The calculated charge for all atoms were shown in Figure 6. It is well-known that
3
the Mulliken charges confirm the hydrogen bonding in the molecular structure of compound.
2
.2.7 Thermodynamic properties
In order to determine thermodynamical properties of the compound (1), the standard
thermodynamic functions, entropy ( ꢀꢁ ) heat capacity (ꢂꢁ ) and enthalpy (ꢅ ) based on the
ꢁ
ꢃꢄꢀ
ꢀ
vibrational analysis at B3LYP/6-311G(d,p) level and statistical thermodynamics for the compound
1) were obtained. The entropies, standard heat capacities, and enthalpies increased at any
(
temperature from 50 K to 900 K since increasing temperature causes an increase in the intensity of
the molecular vibration and the populations of the excited vibrational states. Based on the calculated
data of the compound (1), the correlation equations between the thermodynamic properties and

temperature T had been obtained which can be used for the further studies of the compound (1)
Figure 7).
(
ꢁ
ꢆꢇꢈꢉꢊꢋꢌꢍ ꢏꢐꢑꢒꢓꢉꢔꢍꢏꢒꢑꢓꢕꢉꢒꢇꢖꢗ ꢆꢐꢑꢘꢘꢙꢔꢚꢇꢇꢇꢇꢇꢇꢇꢇꢇꢇꢇꢇꢇꢇꢖꢔꢚ
ꢎ
ꢎ
ꢀ
ꢁ
ꢎ
ꢎ
ꢂ
ꢆꢇꢈꢐꢑꢐꢐꢐꢔꢍ ꢏꢐꢑꢒꢙꢓꢒꢍꢈꢐꢑꢛꢛꢔꢓꢇꢖꢗ ꢆꢐꢑꢘꢘꢕꢓꢚꢇꢇꢇꢇꢇꢇꢇꢖꢕꢚ
ꢃꢄꢀ
ꢁ
ꢎ
ꢎ
ꢅ ꢆꢐꢑꢐꢐꢐꢔꢍ ꢏꢐꢑꢐꢕꢓꢒꢍꢏꢙꢑꢙꢓꢐꢓꢇꢖꢗ ꢆꢐꢑꢘꢘꢔꢕꢚꢇꢇꢇꢇꢇꢇꢇꢇꢇꢇꢇꢇꢇꢖꢒꢚ
ꢀ
2
.3 Molecular docking studies
The molecular docking of compound (1) was done with two target proteins, COX1(1EQG)
28] and COX2 (1CX2) [29] downloaded from the protein data bank. The reference ligands were
[
used to set-up the binding pocket before compound (1) was docked into the target protein. For,
COX1, Ibuprofen which is the reference ligand in the co-crystallized protein gave the parameter used
for the binding pocket where 25.995, 34.157 and 200.016 for the X, Y and Z axes within a radius of
1
6 Å while for COX-2 which had a selective inhibitor, SC 558 has its reference ligand in the co-
crystallized protein gave rise to 24.306, 22.187 and 14.462 for the X, Y and Z axes within a radius of
1
5 Å as the parameters for its binding pocket.
Auto Dock Vina [30], which is a widely used molecular docking program combines both
knowledge based and empirical scoring functions and whose docking score corresponds to binding
energy in kcal/mol was used for the docking analysis. Density functional theory (DFT) method with
the hybrid B3LYP employing the 6-311G(d,p) basis set, that has been shown to give quite
satisfactory geometry results for organic systems [31, 32] using the Gaussian 09 package [21] was
carried out to optimize the structure of compound (1). The optimized compound (1) structure (Fig. 1)
was used as the input ligand in all the docking calculations. The docking result gave rise to 9
different poses (Tables 2 and 3). The first pose which had 0 has both the RMSD (Root Mean Square
Distance) of its upper and lower boundaries was selected to determine the binding modes and
molecular interactions (Figs 8 and 9) and this pose also had the highest binding affinity of -8.7
kcal/mol and -8.1 kcal/mol for both COX-1 and COX-2. Fig.8a shows the binding mode of PD in
COX-1 with ibuprofen the reference ligand highlighted in yellow. The molecular interactions (Fig.
8
b) of compound (1) in the binding pockets include hydrogen bonding with ARG 120, and alkyl
bonding with PHE 518, ILE 523, VAL 349, ALA 527, LEU 531, LEU 359, VAL 116 and LEU 93.
For COX-2, the binding mode (Fig. 9a) shows SC-558 highlighted in yellow. The molecular
interactions (Fig 9b) of compound (1) in the binding pocket includes, hydrogen bonding with MET
5
1
22, carbon-hydrogen bonding with ALA 527, VAL 349 and SER 530; pi-cation bonding with ARG
20; pi-pi stacking with TRY 355 and GLY 526 and alkyl bonding with HIS 90, VAL 523, PHE 518,

LEU 351 and LEU 531. It is worthy to note that there are some common amino acids which
compound (1) interacts with in both COX-1 and COX-2. These includes; ARG 120 and VAL 349.
This may mean that compound (1) is a potential inhibitor for both COX-1 and COX-2.
Docking studies was also conducted with 5-LOX, AChE and BuChE (supporting
information). Compound (1) exhibited the following binding affinities values for all three target
-1
proteins respectively -9.4, -9.1 and -10.5 kcalmol for 5-LOX, AChE and BuChE respectively.
Compound (1) showed highest binding affinity with BuChE. (Supporting Information)
2
2
.4 Biological Activities
.4.1 Anti-inflammatory and anti-ulcer activities
Percent inhibitions of edema by the ibuprofen, sulfanilamide and the compound (1) at 50 mg
-1
kg dose were calculated. These result (Table 4) indicated a substantially enhanced anti-
inflammatory activity of compound 1 compared to ibuprofen and sulphanilamide. Similar results has
been obtained for anti-ulcer test where compound 1 showed substantially reduced ulceration
compared to ibuprofen and sulphanilamide (Table 4). Experiments had also been carried out using
ibuprofen and sulphanilamide at half dose as a control experiment. Again compound 1 was found to
be more active compared to ibuprofen and sulphanilamide at half dose. The main cause of this
enhanced anti-inflammatory and anti-ulcer activity may be due to the presence of polar groups
(sulphonyl and amide) in middle and hydrophobic group (p-isobutyl and benzene) at one side of
compound 1.
2
2
.4.3 Enzyme inhibition studies
.4.3.1 In vitro 5-lipoxygenase (5-LOX) inhibition assay
In vitro anti-inflammatory activity of the compound (1) was evaluated by inhibition of 5-
LOX as this enzyme is involved in an alternative pathway for processing of arachidonic acid
resulting in increased production of pro-inflammatory and gastrotoxic leukotrienes [33]. Compound
(1) exhibited higher inhibition (48.4%) than ibuprofen (42.5%) suggesting that compound (1)
possesses improved activity. Experiments had also been carried out using ibuprofen and
sulphanilamide at half dose as a control experiment. Again compound 1 was found to be more active
compared to ibuprofen and sulphanilamide at half dose. The same results has also been proved by
docking studies (supporting Information). Main cause of this activity is the presence of sulphonyl
and amide group in the vicinity of ARG246 and ARG 370 which provide hydrogen bonding
interactions. Presence of p-isobutyl group provides the cruicial pi-alkyl interactions with Val361,

Leu281 and phe286 within the pocket of enzyme. All these interactions together with others (Figure
, supporting information) are responsible for better binding affinity with compound 1 compared
with ibuprofen.
4
2
.4.3.2 In vitro acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) inhibition
assays
Acetylcholine has been shown to exert an anti-inflammatory function by down-modulating
the expression of pro-inflammatory cytokines [34]. Thus the expression of AChE, being responsible
for acetylcholine hydrolysis, is modulated in inflammation. Generally the BuChE activities resemble
those of AChE, therefore, the compounds under investigation were tested for their inhibitory activity
against these enzymes. It was found (Table 5) that the inhibitory activity of the compound (1) was
substantially less than that of ibuprofen against AChE and almost two times higher against BuChE
(Table 6). This result suggests partially selective inhibition of BuChE by the compound (1) as
compared with ibuprofen. Experiments had also been carried out using ibuprofen and sulphanilamide
at half dose as a control experiment. Again compound 1 was found to be more active compared to
ibuprofen and sulphanilamide at half dose. The same results has also been proved by docking studies
which shows highest affinity of compound 1 with BuChE (supporting information). By comparing
the experimental data and molecular docking studies structure-activity relationship can be deduced.
Here again the sulphonyl, amide and p-isobutuyl groups of compound 1 fit into the right sized pocket
of enzymes AChE and BuChE compared with ibuprofen. His381, Arg525 and Ala397 provides
important pi-alkyl interactions in the case of AChE (Figure 5, Supporting Information) while
phe526, try396 asn229 groups of BuChE interacts with p-isobutuyl, benzene and sulphonyl groups
of compound 1 via pi-alkyl, pi-pi and hydrogen bonding respectively (Figure 6, Supporting
Information).
2
.4.4 Free radical scavenging activities
The free radical scavenging activity of the compound (1) was found (Table 7) to be nearly
double than that of ibuprofen; this also suggests that the compound (1) should exhibit enhanced anti-
inflammatory activity by suppressing the effect of reactive oxygen species.
2
.4.5 Toxicity study
The LD (oral, rats) values were found to be sulfanilamide: 3500 mg kg , ibuprofen: 630 mg kg
-1
-
5
0
1
-1
and compound (1): 4050 mg kg . These results clearly indicate that the compound (1) safer than
the parent drugs.

3
. Conclusion
NSAIDs containing carboxylic groups can be covalently coupled with anti-infectives containing
amino groups to produce mutual prodrugs with dual activity and improved toxicity profile. A novel
compound (1), a mutual prodrug of ibuprofen and sulfanilamide, has been synthesized. Compound
(1) geometry was optimized using density functional theory (DFT/B3LYP) method with the 6-
3
11G(d, p) basis sets in ground state. Calculated geometric parameters (bond lengths, bond angles,
torsion angles), vibrational assignments, chemical shifts and thermodynamics are in good agreement
with the experimental data. Molecule was subjected to Comparative AutoDock study with
cyclooxygenase enzymes (COX-1 and COX-2) involving docking for possible selectivity of
compound (1) within the two Cox enzymes. The highest binding affinities of -8.7 Kcal/mol and -8.1
Kcal/mol has been obtained for COX-1 and COX-2 enzymes respectively. Compound (1) was found
to have enhanced anti-inflammatory, anti-ulcer and free radical scavenging activities when compared
with the parent drugs suggesting that the compound (1) is more active than the parent drugs while
LD of compound (1) is higher than parent drug i.e. ibuprofen and sulphanilamide.
5
0
Acknowledgement
We acknowledge university of Education, Lahore, Pakistan to provide funding and facilities to
accomplish this project.
4
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O
O
Oxalyl Chloride
Cl
OH
5o
C
(A)
NH2
O
S
H2N
O
O
NH2
HN
S
O
O
(1)
Scheme 1. Synthesis of mutual prodrug of ibuprofen and sulfanilamide (1).
Figure 1: Optimized structure of compound (1)

4000
3000
2000
1000
-
1
Wave Number (cm )
Figure 2: Theoretical FT-IR Spectrum of compound (1)
Figure 3: Correlation graphics of calculated and experimental chemical shifts of compound (1)
Figure 4: The HOMO and LUMO molecular orbital diagram

Figure 5: Molecular electrostatic potential (MEP) map of compound (1) in gas phase.
Figure 6: The Mullikan charges diagram of compound (1)
Figure 7: Thermodynamic Properties of compound (1)

Figure 8. The binding mode(A)* and molecular interactions(B) of the title compound in COX2
PDB ID: 1EQG) *the native ligand is in yellow
(
Figure 9: The binding mode(A)* and molecular interactions(B) of the title compound in COX2
PDB ID: 1CX2) *the native ligand is in yellow
(

Table 1: The calculated frontier orbital energies, electronegativity, hardness, and softness of
compound using B3LYP/6-311G(d, p) level.
Parameters
EHOMO
ELUMO
B3LYP/6-311G(d, p)
-6.3085
-1.1793
I (eV)
6.3085
A (eV)
χ (eV)
1.1793
3.7437
η (eV)
2.5646
S (eV)
0.1949
Table 2. Molecular docking results of compound (1) in COX-1
Binding
Poses
Affinity rmsd/ub rmsd/lb
(
kcal/mol)
-8.7
-8.6
-8.6
-8.4
-8
1
2
3
4
5
6
7
8
9
0
0
2.107
9.115
3.221
8.912
3.286
10.319
9.024
7.349
1.553
3.223
2.62
4.505
2.406
6.734
6.944
4.156
-8
-7.9
-7.9
-7.9
Table 3. Molecular docking results of compound (1) in COX-2
Binding
Poses
Affinity rmsd/ub rmsd/lb
kcal/mol)
(
1
2
3
4
5
6
7
8
9
-8.1
-8
-7.3
-7.3
-7
-6.9
-6.9
-6.6
-6.5
0
0
2.267
5.467
5.816
14.335
14.497
5.022
14.748
13.941
1.095
2.838
2.61
12.809
11.266
1.894
12.79
12.362

Table 4: Anti-inflammatory and Anti-Ulcer assay
Anti-
Inflammatory
Anti-Ulcer
Ulcer
Compound
%
%
Inhibition
Area
Ibuprofen
Sulphanilamide
45±0.14
10±0.07
35±0.18
10±0.09
20±0.12
8±0.04
5
0%Ibuprofen+50%Sulphanilamide 23.23±0.11
Compound 1
75±0.16
Table 5: Acetylcholinesterase assay
Concentration of
the Solution used
in Assay (mM)
0.5
Compound
% Inhibition IC50 (µ mol)
Ibuprofen
Sulphanilamide
5
61.19±0.04
10.19±0.05
40.23±0.07
47.21±0.11
235.11±0.17
145.24±0.13
------
0.5
0%Ibuprofen+50%Sulphanilamide 0.5
Compound 1
0.5
< 600
Table 6: Butyrylcholinesterase assay
Concentration of
the Solution used
in Assay (mM)
0.5
Compound
% Inhibition IC50 (µ mol)
Ibuprofen
Sulphanilamide
5
35.51±0.15
5.19±0.07
21.68±0.06
65.88±0.36
< 600
138.79±0.09
------
0.5
0%Ibuprofen+50%Sulphanilamide 0.5
Compound 1
0.5
214.41±0.05
Table 7: DPPH activity
Compound Concentration of
the solution used in (%)
assay (mM)
Inhibition
IC50
(μmol)
Ibuprofen
11.33±0.11 <500
0
.5
Quercetin
0.25
93.21±0.97 16.96±0.14