Synthesis and urease inhibitory potential of benzophenone sulfonamide hybrid in vitro and in silico

Article abstract of DOI:10.1016/j.bmc.2019.01.043

This study deals with the synthesis of benzophenone sulfonamides hybrids (1–31) and screening against urease enzyme in vitro. Studies showed that several synthetic compounds were found to have good urease enzyme inhibitory activity. Compounds 1 (N′-((4′-h

Full text of DOI:10.1016/j.bmc.2019.01.043

Accepted Manuscript
Synthesis and urease inhibitory potential of benzophenone sulfonamide hybrid
in vitro and in silico
Arshia, Farida Begum, Noor Barak Almandil, Muhammad Arif Lodhi, Khalid
Mohammed Khan, Abdul Hameed, Shahnaz Perveen
PII:
S0968-0896(18)32153-9
https://doi.org/10.1016/j.bmc.2019.01.043
BMC 14735
DOI:
Reference:
Bioorganic & Medicinal Chemistry
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Received Date:
Revised Date:
Accepted Date:
24 December 2018
29 January 2019
31 January 2019
Please cite this article as: Arshia, Begum, F., Almandil, N.B., Lodhi, M.A., Khan, K.M., Hameed, A., Perveen, S.,
Synthesis and urease inhibitory potential of benzophenone sulfonamide hybrid in vitro and in silico, Bioorganic &
Medicinal Chemistry (2019), doi: https://doi.org/10.1016/j.bmc.2019.01.043
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Synthesis and urease inhibitory potential of benzophenone sulfonamide hybrid in vitro and
in silico
Arshia,a Farida Begum,b Noor Barak Almandil,c Muhammad Arif Lodhi,b Khalid Mohammed
Khan,a,c Abdul Hameeda, Shahnaz Perveen,d
aH. E. J. Research Institute of Chemistry, International Center for Chemical and Biological
Sciences, University of Karachi, Karachi-75270, Pakistan
bAbdul Wali Khan University Mardan, Khyber Pakhtunkhwa 23200, Pakistan
cDepartment of Clinical Pharmacy, Institute for Research and Medical Consultations (IRMC),
Imam Abdulrahman Bin Faisal University, P.O. Box 31441, Dammam, Saudi Arabia
dPCSIR Laboratories Complex, Shahrah-e-Dr. Salimuzzaman Siddiqui, Karachi-75280, Pakistan
Abstract: This study deals with the synthesis of benzophenone sulfonamides hybrids (1-31) and
screening against urease enzyme in vitro. Studies showed that several synthetic compounds were
found to have good urease enzyme inhibitory activity. Compounds 1 (N'-((4'-
Hydroxyphenyl)(phenyl)methylene)-4''-nitrobenzenesulfonohydrazide),
Hydroxyphenyl)(phenyl)methylene)-3''-nitrobenzenesulfonohydrazide),
Hydroxyphenyl)(phenyl)methylene)-4''-methoxybenzenesulfonohydrazide),
2
3
(N'-((4'-
(N'-((4'-
(3'',5''-
4
Dichloro-2''-hydroxy-N'-((4'-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide),
6 (2'',4''-Dichloro-N'-((4'-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide), 8
(5-(Dimethylamino)-N'-((4-hydroxyphenyl)(phenyl)methylene)naphthalene-1-sulfono
hydrazide),
10
(2''-Chloro-N'-((4'-
12 (N'-((4'-
hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide),
Hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide) have found to be potently
active having an IC50 value in the range of 3.90-17.99 µM. These compounds showed superior
activity than standard acetohydroxamic acid (IC50 = 29.20 ± 1.01 µM). Moreover, in silico
studies on most active compounds were also performed to understand the binding interaction of
most active compounds with active sites of urease enzyme. Structures of all the synthetic
1
compounds were elucidated by H-NMR, 13C-NMR, EI-MS and FAB-MS spectroscopic
techniques.
Keywords: Benzophenone sulfonamide hybrid; urease; in vitro; in silico.
*Corresponding Authors: khalid.khan@iccs.edu, drkhalidhej@gmail.com, Tel.: +922134824910, Fax:
+922134819018
1

1. Introduction
Sulfonamide chemically containing sulfamoyl (SO2NH) group is derivatives of amide. The first
sulfonamide drug identified in 1932 was the prontosil and used as antibacterial agent [1].
Sulfonamides executed as competitive inhibitors due to resemblance with PABA (p-amino
benzoic acid) and hence, block the synthesis of folic acid thus ultimately block the growth of
bacteria [2]. Sulfonamide is biologically significant class of compound, because it is well
absorbed orally and excreted in urine. Thus, it has less toxicity, increased reactivity, and cost
effective [3] molecues. Sulfonamides are mainly used as antibacterial drugs [4] but they also
have anticancer [5], antiinflammatory [6], antiviral [7], antiobesity [8], and carbonic anhydrase
inhibitory activities [9]. Sulfonamide synthesis takes place by nucleophilic substitution of NH
group to sulfonyl chloride. Sulfonamide functionality has also found application in food
additives and in animal husbandry [10].
Benzophenone also called as benzoyl benzene structurally has two phenyl group joined together
through a carbonyl group. Benzopheonone nucleus chemically have nature of secondary
metabolite i.e. polyketide. Naturally benzophenone molecules found in Garcinia
cochinchinensis, and Clusiacea family and it can also be obtained synthetically. The ability of
benzophenones as chemotherapeutic agent especially as anticancer [11], antimicrobial [12],
antiHIV [13], antiinflammatory [14], antioxidant and urease inhibitory activity is well
documented. Benzophenone nucleus also has importance in field of agriculture as one of the
fungicide i.e. fluomorph also have benzophenone as key skeleton.
Urease also known as urease aminohydrolase, is a nickel containing metaloenzyme have enzyme
commission number of (EC 3.5.1.5). Urease enzyme is distributed from prokaryotes to
eukaryotes [15]. Main function of urease involves degradation of urea into carbon dioxide and
ammonia [16]. In human, there is no any urease enzyme is present but they form urea which is
the end product of protein metabolism and normally it excreted from body in the form of urine.
Since, if human is infected from bacteria such as with Helicobacter pyroli which contains
virulence factor i.e. urease enzyme. Urease enzyme is seem to be presence in variety of
pathogenic bacteria. It plays a major role in colonization of a host organism and in protection of
bacterial cells in host tissues therefore, urease has toxic effect on human [17]. Urease enzyme in
pathogeneic bacteria degraded human urea into carbon dioxide and ammonia as a results of
2

which human defence mechanism releases increased amount of stomach acid which ultimately
causes gastric and dueodenal ulcers [18].
To overcome the urease activity of pathogenic bacteria, our research group has been engaged in
designing small, cost effective, and less toxic urease inhibitor since two decades [19-27]. Since,
sulfonamide molecules showed good antibacterial and urease inhibitory activity [28,29]. In
addition, fine urease inhibitory activity of benozophenone derivative is also previously reported
by our research group [30] (Figure-1). Therefore, in this study, we have synthesized and
characterized thirty-one (31) hybrids of benzophenone sulfonamide and screened them against
urease enzyme in vitro. In silico study has also been performed to understand binding mode of
active compound against urease enzyme. To the best of our knowledge compounds 13, 24, 27
and 31 are previously known [31], while remaining compounds are new.
3

Figure-1: Rationale of current sudy
4

2
Results and discussion
Chemistry
2.1
Benzophenone sulfonamide hybrids have been synthesized by the reaction of benzophenone and
4′-hydroxy benzophenone hydrazones with differently substituted sulfonyl chloride, respectively.
Synthesis has been performed by stirring a reaction mixture at room temperature in the presence
of pyridine acting as base as well as solvent which results in precipitation. Reaction was also
performed by taking acetonitrile as solvent and pyridine as a base but no results were obtained.
Chemical structures of these synthetic analogues were characterized by 1H-NMR, 13C-NMR, EI-
MS and FAB-MS techniques Table-1.
Scheme-1: Synthesis of benzophenone sulfonamide hybrid 1-31
2.2
Spectroscopic
studies
of
active
compound
N'-((4'-
hydroxyphenyl)(phenyl)methylene)-4''-methoxybenzenesulfonohydrazide (3)
Active compound 3 is randomly selected for describing detailed characterizing of the structure of
the synthetic benzophenone sulfonamide. For understanding hydrocarbon skeleton NMR spectra
of compound 3 i.e. proton and carbon spectra were run in deuterated dimethyl sulfoxide on a
Bruker Avance AM 300 MHz instrument.
1
In H-NMR spectrum, the most downfield signals at H 10.23 showed the presence of SO2NH
group in the molecule. The phenolic proton resonated at H 9.85. In aryl region the most
downfielded protons resonate as doublet at H 7.85 (J2,3/6,5 for H-2, H-6). Protons 3, 4, 5 resonate
as triplet atH 7.72 (J3(2,4)/4(3,5)/5(4,6) for H-3, H-4, H-5). In aromatic region of proton NMR
spectrum, an aryl ortho protons resonate as doublet at H 7.24 (J2′′,3′′/6′′,5′′ for H-2′′, H-6′′). While
the H-3′′, H-5′′ protons neighbour to methoxy group also appear as doublet at H 7.14 (J3′′,2′′/5′′,6′′
for H-3′′, H-5′′). The most upfield signals in aromatic region is assigned to 4′-substituted hydroxyl
5

benzophenone part, these upfield protons of aromatic region appeared as doublets at H 7.05
(J2′,3′/6′,5′ for H-2′, H-6′) and H 6.88 (J3′,2′/5′,6′ for H-3′, H-5′) (Figure-2).
Figure-2: 1H-NMR analysis of compound 3.
Broad band spectrum of compound 3 showed total of 14 carbons present in the structure. Out of
which there are 6 quaternary, 7 methines and 1 methoxy carbons. While from DEPT-135, it was
also proven that there is no any methylene group in the structure. Iminic carbon showed most
downfield signal at C 162.5. In aromatic region the most downfield signal was supposed to be
C-1′′ at C 158.3. Aryl aromatic carbon attached to methoxy group that is C-4′′ resonate at C
154.5. The two carbons (C-3′, C-5′) of benzophenone resonated at the most upfield region of C
115.4, this is due to the mesomeric effect of the adjacent hydroxy group. All other aromatic
carbons resonate at their usual aromatic range (Figure-3).
Figure-3: 13C-NMR analysis of compound 3.
FAB-MS technique was used for determining mass of compound 3 which gave mass m/z 383
[M+H]+.
6

Figure-4: General structure of synthetic benzophenone sulfonamide
Structure-Activity Relationships (SAR)
2.3
Thirty-one (31) benzophenone sulfonamides were synthesized and screened for urease inhibition
activity. Enzyme inhibition studies suggested that urease inhibition activity of this class of
compounds not only depend on benzophenone part but also at different substitution of alkyl/aryl
sulfonyl part (Figure-4). The limited review at structure-activity relationship reveals that
hydroxylated benzophenone part has showed good inhibitory activity. Moreover, amongst
hydroxylated benzophenone part various substituents at alkyl/aryl sulfonyl part responsible for
varying degree of activity.
Amongst these synthetic compounds, twelve compounds have hydroxyl group at 4′-position of
benzophenone part while nineteen compounds were without hydroxyl group at benzophenone
part. Amongst all synthetic compounds, seventeen compounds showed good to weak inhibitory
activity with IC50 values of 3.90-71.63 μM. Most active compound 1 (IC50 = 3.90 ± 0.81 μM)
found to be about 10-fold more active than acetohydroxamic acid (IC50 = 29.20 ± 1.01 μM)
which was used as the standard and a detailed SAR is discussed in forthcoming paragraphs.
As mentioned earlier, compound 1 having hydroxyl group at 4′- site of benzophenone and nitro
group at 4′′- position of aryl sulfonyl part was found to be most active compound of series,
however, switching nitro group to 3′′- position of aryl sulfonyl part as in compound 2 (IC50
=
13.73 ± 0.01 μM) a decreased activity was observed. Removal of hydroxy group from
benzophenone part and shifting of nitro group to 2′′/3′′/4′′-positions, respectively, resulted in
inactive compounds 19, 20, and 21 (Figure-5) which indicates that the nitro group is only
effective when it is in combination of hydroxyl group at benzophenone part.
7

Figure-5: SAR of nitro substituted aryl sulfonyl part
However, nitro group in combination with chloro group at aryl sulfonyl part in absence of
hydroxyl group at benzophenone part, as in compounds 22 and 25 showed no any activity.
Nevertheles, compound 24 (IC50 = 68.9 9 ± 0.71 μM) having dinitro substituent at 2′′,4′′-
positions of aryl sulfonyl benzophenone, and compound 23 (IC50 = 68.99 ± 0.65 μM) having
methyl group at 2′′- position and nitro group at 5′′-position in combination of non-hydroxylated
benzophenone was found to be weakly active (Figure-6).
8

Figure-6: SAR of disubstituted i.e. nitro and chloro substituted aryl sulfonyl part
In hydroxylated benzophenone molecules, compound 3 (IC50 = 14.51 ± 1.14 μM) having
methoxy group at 4′′-position of aryl sulfonyl part showed good inhibitory activity. However,
non-hydroxylated compounds 27 and 28 having methoxy groups at 4′′-position and at 2′′ and 4′′-
positions, respectively, were found to be inactive (Figure-7).
Figure-7: SAR of methoxy substituted aryl sulfonyl part
Compound 4 (IC50 = 4.97 ± 0.31 μM) having hydroxylated benzophenone part and in aryl
sulfonyl part hydroxyl group at 2′′-position and chloro group at 3′′,5′′-positions was found to be
second most active compound of the series. Nevertheles, compound without any hydroxyl group
substitution at benzophenone as in compound 26 (IC50 = 71.63 ± 0.81 μM) showed weak
inhibitory activity. Shifting of chloro group at 2′′,4′′-positions of aryl sulfonyl part and hydroxyl
group at 4′-position makes compound 6 (IC50 = 17.99 ± 0.61 μM) showed good but relatively
weak inhibitory activity than compound 4. However, analogs having no hydroxyl group at
benzophenone part, compound 18 was found to be completely inactive. Presence of only one
9

chloro group at 2′′-position and hydroxyl group at 4′-positions as in compound 10 (IC50 = 16.91 ±
2.10 μM) showed comparable activity to compound 6. However, removal of hydroxyl substituent
from benzophenone part and chloro group at 2′′-position of aryl sulfonyl part as in compound 14
made it completely inactive. However, shifting of chloro group at 2′′-position without hydroxyl
group at benzophenone part as in compound 16 (IC50 = 66.10 ± 0.11 μM) exhibited weak urease
inhibitory activity. Chloro group at 4′′-position and methyl group at 2′′, 5′′-positions of aryl
sulfonyl part without hydroxyl group at benzophenone part as in compound 15 made it
completely inactive. Trichloro substituted aryl sulfonyl part with chloro group at 2′′,4′′,5′′-
positions and hydroxyl group at 4′-position of aryl sulfonyl part as in compound 9 (IC50 = 57.88
± 0.71 μM) showed weak inhibitory activity. Compound 17 with trichloro substituent at same
positions of aryl sulfonyl part as in compound 9 but without hydroxyl group at benzophenone
part made it completely inactive (Figure-8).
10

Figure-8: SAR of chloro, hydroxyl and methyl substituted aryl sulfonyl part
Compound 7 (IC50 = 55.04 ± 0.54 μM) with hydroxyl group at benzophenone part and
chloroethane residue at aryl sulfonyl part made it weakly active. Only methyl substituent instead
of aryl sulfonyl part as in compound 11 (IC50 = 43.39 ± 0.41 μM) demonstrated weak inhibitory
activity. Compound 31 with no any substituent at benzophenone part and have butyl chain
instead of aryl position made it completely inactive (Figure-9).
11

Figure-9: SAR of alkyl substituted aryl sulfonyl part
Benzene ring at aryl sulfonyl part without hydroxyl group substituent at benzophenone part as in
compound 13 showed no any activity. Exchanging of benzene with naphthalene ring as in
compound 29 also is ineffective. However, N,N-dimethylnaphthalene as aryl sulfonyl part as in
compound 30 (IC50 = 72.19 ± 0.60 μM) found to be weakly active. Whereas, compound 8 (IC50 =
5.70 ± 0.19 μM) with hydroxylated benzophenone has same other functionalities of compound
30 was found to be third most active compound of the series (Figure-10).
Figure-10: SAR of benzene and N,N-dimethyl naphthyl substituted aryl sulfonyl part
12

Table-1: Synthetic compound 1-31 with urease inhibitory activities in vitro
Compound No.
1
R
R′
IC50 ± SEMa [μM]
OH
3.90 ± 0.81
OH
OH
13.73 ± 0.01
14.51 ± 1.14
2
3
OH
OH
4.97 ± 0.31
4
5
31.20 ± 0.16
OH
OH
17.99 ± 0.61
55.04 ± 0.54
6
7
OH
OH
5.70 ± 0.19
8
9
57.88 ± 0.71
OH
OH
16.91 ± 2.10
43.39 ± 0.41
10
11
-CH3
13

OH
H
16.30±1.10
NAb
12
13
H
H
NAb
NAb
14
15
H
H
H
66.10 ± 0.11
NAb
16
17
18
NAb
H
H
NAb
NAb
19
20
H
NAb
21
14

H
H
NAb
22
23
68.99 ± 0.65
H
68.9 9± 0.71
24
H
H
NAb
25
26
71.63 ± 0.81
H
NAb
27
H
H
NAb
NAb
28
29
H
H
72.19 ± 0.60
30
31
NAb
29.20 ± 1.01
Acetohydroxamic acid
SEMa = Standard Error of the Mean; NA.b = Not Active
15

2.4
Computational studies:
In order to under the binding mode of these newly synthesized compounds, molecular docking
was performed. From the docking conformation of the compounds, it was revealed that
compound 1 (IC50 = 3.90 ± 0.81, docking score = -21.4737), showed arene-cation interaction
with catalytic residue His323 and also established linkages with both nickel of the active site of
the urease enzyme as shown in Figure 11(a). The presence of -NO2 group at para position made
it most active compound in the series. The second most active compound was compound 4 (IC50
= 4.97 ± 0.31, docking score = -21.3977) showed similar interactions, one of the oxygen of the
phenyl moiety of ligand contact with nickel and second phenyl moiety formed arene-cation
linkage with His323 as shown in(Figure 11(b)).Compound 1 and 4, both exhibited the excellent
biological activities against the urease enzyme, however, if compared compound 1 with the
compound 4 having slight difference in the activities, due to the presence of -Cl group on ortho
position, -OH and -Clgroups at meta positions which made compound 4 the second most active
compound and in the series. The third most active compound 8 (IC50 = 5.70±0.19, docking score
= -21.2256) showed one arene-cation contact with the His323 residue of the binding pocket of
the urease enzyme. The oxygen of phenyl moiety of sulfonohydrazide showed hydrogen bonding
with His275 and also established contact with nickel (Figure-11c). The fourth one active
compound is 2 (IC50 = 6.10 ± 0.01, docking score = -20.7057) which showed one arene-cation
interaction with the catalytic residue His 323 of the enzyme. The oxygen of the phenyl moiety
mediates interaction with both nickel as shown in (Figure-11d). Compound 3 showed two arene-
cation interactions, one with His323 and second with nickel, the second nickel involved in
interaction with His249, His222, and KCX220 which established contact with oxygen of the
phenyl residue of the benzenesulfonohydrazide of the active site of the urease enzyme shown in
(Figure-11e). The presence of -OMe group at meta position made it fifth most active compound
of the series. Compound 6 (IC50 = 17.99 ± 0.61, docking score = -20.544), oxygen of the phenyl
moiety interact with both nickel. The ligand also showed arene action cationcontact with His323
of the active site of the urease (Figure-11f). Compound 6 have -Cl groups at ortho and para
positions which made it sixth most active compound of the series. The 7th most active compound
12 (IC50 = 16.30 ± 1.10, docking score = -20.2789) mediate arene-arene interaction with catalytic
residue His323 and also phenyl oxygen contact with nickel which subsequently showed binding
with His222, His249 and KXC220 (modified lysine) (Figure-11g). 8th most active compound 10
16

(IC50 = 16.91 ± 2.10, docking score = -20.2587) exhibited similar interactions as shown by
compound 12 (Figure-11h). Both compounds showed excellent biological activities they have
slight difference in their structures at position ortho (-Cl in compound 10) while no any
substitution at aryl sulfonyl part, make compound 12.
Compounds 1-4, 6, 8, 10, 12 of the series showed good inhibitory activities as compared to the
standard acetohydroxamic acid compound (IC50 = 29.20 ± 1.01 μM, docking score = -5.8103)
and these compounds of the series are well fitted in the active site of the urease enzyme. The
control, acetohydroxamic acid makes one hydrogen bond with the active site residues Asp224.
The basic catalytic residues His323 and Arg339 of the urease made two hydrogen bonds with
two oxygen moieties of the standard compound (Figure-11g). Overall a good correlation was
observed between the docking study and biological evaluation of active compounds.
a)
b)
17

c)
d)
e)
18

f)
g)
h)
19

Figure-11: Docking conformation of compound 1 (a), compound 4 (b) 8 (c), compound 2 (d)
and 3 (e) compound 6 (f), compound 12 (g), compound 10 (h)
a)
Figure-12: Docking conformation acetohydroxamic acid (a) in the active site of urease.
3
Conclusion
This study deals with the synthesis of benzophenone sulfonamides (1-31) and their urease
inhibitory potential. Amongst thirty-one compounds, seventeen (1-12, 16, 23, 24, 26, 30)
demonstrated good to moderate inhibitory potential having an IC50 value range of 3.90-71.63
µM. Compounds 1-4, 6, 8, 10, 12 showed superior activity (IC50 = 3.90 - 17.99 µM) than the
standard acetohydroxamic acid (IC50 = 29.20 ± 1.01). Limited structure-activity relationship
revealed that compounds having hydroxyl group at benzophenone part displayed good inhibitory
activity than non-hydroxyl benzophenone part. However, different substitution at aryl sulfonyl
20

part also responsible for varying degree of activity. Moreover, computational studies of most
active compounds showed good binding interaction between active compound and urease
enzyme. Decisively this study provides some leads compounds as urease inhibitors.
4.
Experimental:
4.1
Material and Methods:
Pyridines, benzophenone hydrazone, 4’-hydroxy benzophenone hydrazone, varyingly substitute
sulfonyl chloride and hexane, were purchased from Sigma-Aldrich, USA and used as received
without purification. Thin layer chromatography (TLC) was performed on pre-coated silica gel
aluminium plates (Kieselgel 60, 254, E. Merck). UV wavelength 254 and 365 nm were used for
TLC visualization. Melting points of all the compounds were determined on Stuart® SMP10
1
melting point apparatus and are uncorrected. H-NMR experiments were performed out on
Bruker Avance AM 300, 400, and 500 MHz machines. EI-MS and FAB-MS was recorded on a
Finnigan MAT-311A (Germany) mass spectrometer. 13C-NMR experiments were carried on
Bruker Avance AM 300 and 400 MHz instruments at 75, 100, and 150 MHz, respectively.
Infrared (IR) spectra were recorded on JASCO IR-A-302 spectrophotometer.
4.2. Urease inhibitory assay
Spectrophotometically urease inhibition assay was performed. For urease inhibition assay 5 μL
of synthetic compound was incubated with 25 μL of urease solution (1 U/well) (250 μL) at 30 °C
for 15 minute. After that, 55 μL substrate urea with 100 mM concentration was added and the
plate was again incubated at 30 °C. After incubation 70 μL of basic reagent (0.5% w/v NaOH and
0.1% NaOCl) and 45 μL of carbolic acid (1% w/v carbolic acid and 0.005% w/v
°
Na2[Fe(CN)5NO]) were added at each well. Again plate was incubated for 50 min at 30 C. Rate
of production ammonia was used for determining urease inhibitory activity by following
Weatherburn method and change in absorbance was monitored at 630 nm on a ELISA plate
reader (Spectra Max M2, Molecular Devices, CA, USA). Acetohydroxamic acid was used as a
standard compound [32].
21

4.3. Methodology
Molecular docking
The molecular docking was performed by using MOE-Dock implemented in the Molecular
Operating Environment (MOE), using the default parameters [33]. The X-ray crystallographic
structure of Bacillus pasteurii (PDB Code: 4UBP) was downloaded from the protein databank
[34]. All molecules of water were removed from the structure of the protein. Then, hydrogen’s
were added, and the energy minimization was carried out using a default force field. 3D structure
of all the compounds was built using the Molecular Builder program implemented in MOE.
Finally, a database was created in which all the ligands were converted into their respective 3D
structures and this database was used as input file MOE-docking. Subsequently, the energy of
compounds present in the database was minimized up to 0.05 Gradient using MMFF94x force
field and add hydrogen to the compounds. The database was docked into the active site of
protein using the Triangular Matching docking method and 10 conformations of each Ligand
protein complex were generated with docking score (S). Each complex was analyzed for
interactions and their 3D images were taken.
General procedures for the synthesis of compound 1-31
The synthesis of benzophenone sulfonamide has been performed by stirring the reaction of
benzophenone hydrazone (1 mmol) and 4′-hydroxy benzophenone hydrazone (1 mmol) with
differently substituted sulfonyl chloride (1 mmol) respectively in 10 mL of pyridine. Stirring of
reaction mixture was carried out at for 24 hours. After completion of reaction, reaction mixture
was poured in ice cold water. Which results in precipitation, these precipitate were filter dry and
then washed with hexane. Chemical structures of these synthetic analogues were characterized
by 1H-NMR, 13C-NMR, EI-MS and FAB-MS spectroscopy.
N'-((4'-Hydroxyphenyl)(phenyl)methylene)-4''-nitrobenzenesulfonohydrazide (1)
Yield: 89%; m.p 165-168 °C; Rf: 0.38 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.64 (s, 1H, NH), 9.96 (s, 1H, OH), 8.06 (m, 2H, H-3′′, H-5′′), 7.91 (m, 2H, H-2′′, H-6′′),
7.37 (m, 2H, H-2, H-6), 7.31 (m, 1H, H-4), 7.26(m, 2H, H-3, H-5), 7.11 (d, 2H, J2′,3′/6′,5′ = 8.4 Hz,
H-2′, H-6′), 6.93 (d, 2H, J3′,2′/5′,6′ = 8.7 Hz, H-3′, H-5′); FAB-MS m/z 398 [M+H]+.
22

N'-((4'-Hydroxyphenyl)(phenyl)methylene)-3''-nitrobenzenesulfonohydrazide (2)
Yield: 90%; m.p 170-172 °C; Rf: 0.39 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.71 (d, 1H, J = 78.0 Hz, NH), 9.93 (d, 1H, J = 15.0 Hz, OH), 8.65 (s, 1H, H-2′′). 8.53
(d, 1H, J6′′,5′′ = 8.4 Hz, H-6′′), 8.34 (d, 1H, J4′′,5′′ = 7.8 Hz, H-4′′), 7.97 (t, 1H, J5′′(4′′ 6′′)
= 7.8 Hz, H-
,
5′′), 7.50 (m, 1H, H-4), 7.36 (m, 4H, H-2, H-3, H-5, H-6), 7.21 (m, 2H, H-2′, H-6′), 7.07 (d, 1H,
J2′,3′ = 8.4 Hz, H-3′), 6.86 (d, 1H, J5′,6′ = 8.4 Hz, H-5′); FAB-MS m/z 398 [M+H]+
N'-((4'-Hydroxyphenyl)(phenyl)methylene)-4''-methoxybenzenesulfonohydrazide (3)
Yield: 90%; m.p 168-170 °C; Rf: 0.39 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.23 (s, 1H, NH), 9.85 (s, 1H, OH), 7.85 (d, 2H, J2,3/6,5 = 9.0 Hz, H-2, H-6), 7.36 (t, 3H,
J3(4,5)/4(3,5)/5(4,6)= 8.5 Hz, H-3, H-4, H-5), 7.24 (d, 2H, J2′′,3′′/6′′,5′′ = 9.0 Hz, H-2′′, H-6′′), 7.14 (d, 2H,
J3′′,2′′/5′′,6′′ = 9.0 Hz, H-3′′, H-5′′), 7.05 (d, 2H, J2′,3′/6′,5′ = 8.4 Hz, H-2′, H-6′), 6.88 (d, 2H, J3′,2′/5′,6′
=
13
8.7 Hz, H-3′, H-5′); C-NMR: (75.0 MHz, DMSO-d6): C 162.5 (C=N), 158.3 (C-1′′), 154.5 (C-
4′′), 137.7 (C-3′′, C-5′′), 130.4 (C-1), 129.8 (C-1′), 129.4 (C-2, C-6), 128.1 (C-4) 127.4 (C-2′, C-
6′), 122.8 (C-4′), 115.4 (C-3′, C-5′), 114.0 (C-3, C-5), 55.6 (C-4′′-OCH3); FAB-MS m/z 383
[M+H]+.
3'',5''-Dichloro-2''-hydroxy-N'-((4'-hydroxyphenyl)(phenyl)methylene)benzenesulfono-
hydrazide (4)
Yield: 89%; m.p 178-180 °C; Rf: 0.37 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.09 (s, 1H, NH), 8.59 (d, 1H, J4′′,6′′ = 2.4 Hz, H-4′′), 8.59 (d, 1H, J6′′,4′′ = 2.4 Hz, H-6′′),
7.58 (m, 5H, H-2, H-3, H-4, H-5, H-6), 7.18 (d, 2H, J2′,3′/6′,5′ = 8.4 Hz, H-2′, H-6′), 6.98 (d, 2H,
J3′,2′ = J5′,6′ = 8.7 Hz, H-3′, H-5′); FAB-MS m/z 438 [M+H]+.
2'',5''-Dichloro-N'-((4'-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide (5)
Yield: 83%; m.p 118-120 °C; Rf: 0.37 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.74 (s, 1H, NH), 9.93 (s, 1H, OH), 7.97 (d, 1H, J6′′,3′′ = 1.5Hz, H-6′′), 7.77 (d, 2H, J3′′,4′′
= J4′′,3′′ = 7.5 Hz, H-3′′, H-4′′), 7.36 (m, 2H, H-3, H-5), 7.22 (d, 3H, J2,3/6,5/4,3 = 6.9 Hz, H-2, H-6,
H-4), 7.10 (d, 2H, J2′,3′/6′,5′ = 7.5 Hz, H-2′, H-6′), 6.91 (d, 2H, J3′,2′/5′,6′ = 7.5 Hz, H-3′, H-5′); FAB-
MS m/z 421 [M+H]+.
23

2'',4''-Dichloro-N'-((4'-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide (6)
Yield: 91%; m.p 168-170 °C; Rf: 0.37 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.90 (s, 1H, NH), 7.99 (s, 1H, OH), 7.79 (dd, 1H, J5′′,6′′ = 6.6 Hz, J5′′,3′′ = 2.1 Hz, H-5′′),
7.75 (d, 1H, J6′′,5′′ = 7.5 Hz, H-6′′), 7.55 (m, 3H, H-2, H-6, H-H-3′′), 7.38 (t, 1H, J4(5,6) = H-4),
7.33 (t, 2H, J3(2,4) /5(4,6) = H-3, H-5), 7.27 (m, 2H, H-2′, H-6′), 7.24 (m, 2H, H-3′, H-5′); 13C-NMR:
(100.0 MHz, DMSO-d6): C 155.5 (C=N), 138.0 (C-1′′), 136.8 (C-2′′), 134.3 (C-4′′), 133.7 (C-1′),
132.1 (C-1), 132.0 (C-3, C-5), 130.8 (C-4′), 129.9 (C-2, C-6), 129.8 (C-3′′), 129.6 (C-5′′), 129.0
(C-6′′), 128.6 (C-3′, C-5′), 128.4 (C-4), 127.2 (C-2′, C-6′); FAB-MS m/z 421 [M+H]+.
2''-Chloro-N'-((4'-hydroxyphenyl)(phenyl)methylene)ethanesulfonohydrazide (7)
Yield: 90%; m.p 172-174 °C; Rf: 0.39 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.39 (s, 1H, NH), 9.19 (d, 1H, J = 5.7 Hz, OH), 8.64 (t, 1H, J4(3,5) = 7.5 Hz, H-4), 8.18
(t, 2H, J3(2,4)/5(4,6) = 7.5 Hz, H-3, H-5), 7.55 (m, 2H, H-2, H-6), 7.41 (m, 4H, H-2′, H-3′, H-5′, H-
13
6′), 5.10 (t, 2H, JCH2(CH2) = 6.0 Hz, CH2), 4.19 (t, 2H, JCH2(CH2) = 6.0 Hz, CH2); C-NMR: (75.0
MHz, DMSO-d6): C 155.8 (C=N), 146.1 (C-1′′), 145.5 (C-1′), 136.9 (C-1), 129.9 (C-2, C-6),
129.5 (C-2′, C-6′), 128.3 (C-4), 127.6 (C-4′), 127.6 (C-3, C-5), 54.7 (CH2), 49.2 (CH2); FAB-MS
m/z 339 [M+H]+.
5-(Dimethylamino)-N'-((4-hydroxyphenyl)(phenyl)methylene)naphthalene-1-sulfono-
hydrazide (8)
Yield: 87%; m.p 130-132 °C; Rf: 0.36 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 11.03 (m, 1H, NH), 8.50 (m, 2H, H-3′, H-′), 8.40 (m, 2H, H-2′, H-6′), 7.69 (m, 4H, H-2,
H-6, H-3, H-5), 7.27 (m, 4H, H-4, H-2′′, H-3′′, H-4′′, ), 7.02 (m, 1H, H-6′′), 6.95 (m, 2H, H-7′′, H-
8′′), 2.77 (s, 6H, N(CH3)2); FAB-MS m/z 446 [M+H]+.
2'',4'',5''-Trichloro-N'-((4'-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide (9)
Yield: 89%; m.p 165-167 °C; Rf: 0.35 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.93 (s, 1H, NH), 9.90 (s, 1H, OH), 8.66, (m, 1H, H-3′′), 8.40 (m, 1H, H-6′′), 7.75 (d, 1H,
J4(3,5) = 7.5 Hz, H-4), 7.39, (m, 2H, H-2, H-6), 7.21 (m, 2H, H-3, H-5), 7.09 (d, 2H, J2′,3′/6′,5′ = 8.4
13
Hz, H-2′, H-6′), 6.89 (d, 2H, J3′,2′/5′,6′ = 8.7 Hz, H-3′, H-5′); C-NMR: (150.0 MHz, DMSO-d6):
24

C 148.5 (C=N), 142.6 (C-1′′), 139.3 (C-4′′), 138.6 (C-5′′), 134.2 (C-2′′), 133.1 (C-2, C-6), 133.0
(C-1′), 132.3 (C-4′), 131.5 (C-1), 131.4 (C-3′, C-5′), 131.3 (C-2′, C-6′), 128.2 (C-3′′), 127.3 (C-6′′),
125.3 (C-3, C-5), 122.9, (C-3,C-5); FAB-MS m/z 456 [M+H]+.
2''-Chloro-N'-((4'-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide (10)
Yield: 87%; m.p 130-132 °C; Rf: 0.35 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.33 (s, 1H, NH), 9.92 (s, 1H, OH), 8.05 (d, 1H, J3′′,4′′ = 7.8Hz, H-3′′), 7.69 (m, 2H, H-2,
H-6), 7.62 (m, 1H, H-4), 7.34 (m, 3H, H-3, H-5, H-5′′), 7.19 (d, 2H, J3′′,4′′/6′′,5′′ = 8.4 Hz H-3′′, H-
6′′), 7.10 (d, 2H, J2′,3′/6′,5′ = 8.4 Hz, H-2′, H-6′), 6.93 (d, 2H, J3′,2′/5′,6′ = 8.4 Hz, H-3′, H-5′); FAB-
MS m/z 387 [M+H]+.
N'-((4'-Hydroxyphenyl)(phenyl)methylene)methanesulfonohydrazide (11)
Yield: 89%; m.p 145-147 °C; Rf: 0.37 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 8.68 (s, 1H, NH), 7.67 (s, 1H, OH), 7.65 (m, 2H, H-3, H-5),7.57 (d, 2H, J2,3/6,5 = 6.0 Hz,
H-2, H-6), 7.52 (d, 2H, J2′,3′/6′,5′ = 6.0 Hz, H-2′, H-6′), 7.48 (d, 2H, J3′,2′/5′,6′ = 7.2 Hz, H-3′, H-5′),
6.91 (d, 1H, J4(3,5) = 7.8 Hz, H-4), 2.33 (s, 3H, S-CH3); FAB-MS m/z 291 [M+H]+.
N'-((4'-Hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide (12)
Yield: 86%; m.p 193-195 °C; Rf: 0.38 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.44 (s, 1H, NH), 9.87 (s, 1H, OH), 7.92 (d, 2H, J2,3 = J6,5 = 6.9Hz, H-2, H-6), 7.67 (m,
3H, H-3, H-5, H-4), 7.36 (m, 3H, H-3′′, H-5′′, H-4′′), 7.22 (d, 2H, J2′′,3′′/6′′,5′′ = 6.9 Hz, H-2′′, H-6′′),
7.06 (d, 2H, J2′,3′/6′,5′ = 8.4 Hz, H-2′, H-6′), 6.88 (d, 2H, J3′,2′/5′,6′ = 8.4 Hz, H-3′, H-5′); FAB-MS
m/z 353 [M+H]+.
N'-(Diphenylmethylene)benzenesulfonohydrazide (13)
Yield: 85%; m.p 165-167 °C; Rf: 0.40 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.51(s, 1H, NH), 7.93 (d, 2H, J2,3/J6,5 = 8.4 Hz, H-2, H-6), 7.66 (m, 1H, H-4′′), 7.52 (m,
1H, H-4), 7.44 (m, 1H, H-4′), 7.37 (m, 4H, H-2′′, H-6′′, H-3′′, H-5′′), 7.29 (m, 4H, H-3, H-5, H-3′,
H-5′), 7.22 (m, 2H, H-2′, H-6′); EI-MS: m/z (rel. abund.%), 336 [M]+ (5.8), 283 (95.5), 257
(13.6), 195 (100.0), 180 (38.6), 165 (64.5), 77 (40.9).
25

2''-Chloro-N'-(diphenylmethylene)benzenesulfonohydrazide (14)
Yield: 88%; m.p 120-122 °C; Rf: 0.38 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.57 (s, 1H, NH), 8.06 (d, 1H, J3′′,4′′ = 7.5 Hz, H-3′′), 7.69 (d, 2H, J2,3/6,5 = 8.4Hz, H-2, H-
6), 7.62 (m, 2H, H-3, H-5), 7.56 (d, 2H, J4′ (3′,5′)/4(3,5) = 8.4 Hz, 2H, H-4, H-4′), 7.36 (m, 1H, H-6′′),
7.30 (t, 2H, J4′′(3′′,5′′)/5′′(4′′,6′′) = 8.4 Hz, H-4′′, H-5′′), 7.26 (m, 2H, H-3′, H-5′), 7.17 (d, 2H, , J2′,3′/6′,5′
= 8.4 Hz, H-2′, H-6′); FAB-MS m/z 371 [M+H]+.
4''-Chloro-N'-(diphenylmethylene)-2'',5''-dimethylbenzenesulfonohydrazide (15)
Yield: 88%; m.p 170-172 °C; Rf: 0.39 (ethyl acetate/hexanes, 4:6); 1H-NMR (400 MHz, DMSO-
d6): H 10.64 (s, 1H, NH), 7.87 (s, 1H, H-3′′), 7.52 (m, 4H, H-6′′, H-4, H-4′, H-5), 7.36 (m, 3H, H-
3, H-3′, H-5′), 7.22 (m, 4H, H-2, H-6,H-2′, H-6′), 2.59 (s, 3H, 2′′-CH3), 2.37 (s, 3H, 6′′-CH3); EI-
MS m/z (% rel. abund.): 400 [M+2]+ (14.1), 398 [M]+ (32.6), 360 (67.6), 283 (84.5), 195 (100.0),
165 (91.1), 92 (27.0), 77 (56.9).
4′′-Chloro-N'-(diphenylmethylene)benzenesulfonohydrazide (16)
Yield: 89%; m.p 180-182 °C; Rf: 0.39 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 7.60 (d, 4H, J2,3/6,5/2′′,3′′/6′′,5′′ = 8.4 Hz, H-2, H-6, H-2′′, H-6′′), 7.47 (d, 2H, J3′′,2′′/5′′,6′′ = 8.4
Hz, H-3′′, H-5′′), 7.37 (m, 7H, H-3, H-5, H-2′, H-6′, H-3′, H-5′, H-4), 7.28 (m, 1H, H-4′); 13C-
NMR: (100.0 MHz, DMSO-d6): C 159.0 (C=N), 147.2 (C-1′′), 137.4 (C-4′′), 137.4 (C-1′′), 135.0
(C-1′′), 132.9 (C-1′), 129.9 (C-1), 128.9 (C-2, C-6), 128.8 (C-2′, C-6′), 128.3 (C-2′′, C-6′′), 128.3
(C-3′′, C-5′′), 128.1 (C-4), 128.0 (C-3, C-5), 127.6 (C-3′, C-5′), 127.4 (C-4′); FAB-MS m/z 371
[M+H]+.
2,4,5-Trichloro-N'-(diphenylmethylene)benzenesulfonohydrazide (17)
Yield: 88%; m.p 140-142 °C; Rf: 0.37 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.96 (s, 1H, NH), 8.14 (s, 2H, H-3′′, H-6′′), 7.55 (m, 1H, H-4), 7.47 (m, 1H, H-4′), 7.37
(m, 4H, H-2, H-6, H-2′, H-6′), 7.30 (m, 4H, H-3, H-5, H-3′, H-5′); EI-MS m/z (% rel. abund.):
440[M+2]+ (5.3), 437.0 [M]+ (5.4), 360 (35.6), 283 (61.3), 195 (100.0), 165 (65.5).
26

2'',4''-Dichloro-N'-(diphenylmethylene)benzenesulfonohydrazide (18)
Yield: 90%; m.p 146-148 °C; Rf: 0.36 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.89 (s, 1H, NH), 7.99 (d, 1H, , J3′′,5′′ = 2.1 Hz, H-3′′), 7.78 (m, 1H, H-5′′), 7.55 (m, 2H,
H-4, H-6′′), 7.47 (d, 2H, J2,3/6,5 = 6.9 Hz, H-2, H-6), 7.37 (m, 3H, H-2′, H-6′, H-4′), 7.27 (m, 4H,
H-3, H-5, H-3, H-5); FAB-MS m/z 405 [M+H]+.
N'-(Diphenylmethylene)-2''-nitrobenzenesulfonohydrazide (19)
Yield: 87%; m.p 144-146 °C; Rf: 0.39 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.71 (s, 1H, NH), 8.08 (m, 2H, H-3′′, H-6′′), 7.91 (m, 1H, H-5′′), 7.56 (m, 1H, H-4′′), 7.49
(d, 2H, J2,3/J6,5 = 6.0 Hz, H-2, H-6) 7.37 (m, 4H, H-2, H-6, H-4, H-4′), 7.29 (m, 4H, H-3, H-5, H-
3′, H-5′); EI-MS m/z (% rel. abund.): 381 [M]+ (2.2), 360 (62.3), 283 (100.0), 257, (18.5), 195
(44.3), 180, (41.4), 165 (51.0).
N'-(Diphenylmethylene)-3′′-nitrobenzenesulfonohydrazide (20)
Yield: 92%; m.p 230-232 °C; Rf: 0.40 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 8.32 (m, 4H, NH, H-2′′, H-4′′, H-6′′), 8.19 (m, 3H, H-4, H-4′, H-5′′), 8.02 (d, 4H,
J2,3/6,5/2′,3′/6′,5′ = 7.8 Hz, H-2, H-6, H-2′, H-6′), 7.67 (t, 4H, J3(2,4)/5(6,4)/3′ (2′,4′) 5′ (6′,4′)
= 7.8 Hz, H-3, H-
/
5, H-3′, H-5′); FAB-MS m/z 382 [M+H]+.
N'-(Diphenylmethylene)-4''-nitrobenzenesulfonohydrazide(21)
Yield: 87%; m.p 150-152 °C; Rf: 0.38 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.83 (s, 1H, NH), 8.47 (d, 2H, J3′′,2′′/5′′,6′′ = 9.0 Hz, H-3′′, H-5′′),8.18 (d, 2H, J2′′,3′′/ 6′′,5′′
J
=
8.7 Hz, H-2′′, H-6′′), 7.53 (m, 2H, H-4, H-4′), 7.39 (m, 4H, H-2, H-6, H-2′, H-6′), 7.26 (m, 4H, H-
3, H-5, H-3′, H-5′); EI-MS m/z (% rel. abund.): 381 [M]+ (7.3), 360 (12.9), 283 (21.4), 195,
(100.0), 165, (38.5).
2-Chloro-N'-(diphenylmethylene)-3-nitrobenzenesulfonohydrazide (22)
Yield: 90%; m.p 154-156 °C; Rf: 0.39 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 8.12 (s, 1H, H-NH), 7.86 (dd, 1H, J6′′,5′′ = 6.6 Hz, J6′′,4′′ = 2.1 Hz, H-6′′), 7.74 (d, 1H, J4′′,5′′
27

= 8.4 Hz, H-4′′), 7.47 (d, 2H, J2,3/6,5 = 6.9 Hz, H-2, H-6), 7.37 (m, 5H, H-3, H-5, H-3′, H-5′, H-5′′),
7.29 (m, 4H, H-2′, H-6′, H-4, H-4′); FAB-MS m/z 416 [M+H]+.
N'-(Diphenylmethylene)-2''-methyl-5''-nitrobenzenesulfonohydrazide (23)
Yield: 91%; m.p 160-162 °C; Rf: 0.40 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 11.01 (s, 1H, NH), 8.68 (d, 1H, J6′′,4′′ = 2.4Hz, H-6′′), 8.41 (dd, 1H, J4′′,3′′ = 6.0 Hz, J4′′,6′′
=
2.4 Hz, H-4′′), 7.76 (d, 1H, J3′′,2′′ = 8.4 Hz, H-3′′), 7.54 (m, 3H, H-2, H-4, H-6), 7.37 (m, 3H, H-2′,
H-4′, H-6′), 7.23 (m, 4H, H-3, H-5, H-3′, H-5′), 2.80 (s, 3H, H-2′′-CH3); EI-MS m/z (% rel.
abund.): 395 [M]+ (2.5), 195, (60.1), 165, (100.0), 137, (17.0), 91 (15.5), 77 (13.2).
N'-(Diphenylmethylene)-2'',4''-dinitrobenzenesulfonohydrazide (24)
Yield: 90%; m.p 155-156 °C; Rf: 0.38 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 11.31 (s, 1H, NH), 8.97 (m, 1H, H-3′′), 8.70 (d, 1H, J4(3,5) = 7.5 Hz, H-4), 8.42 (dd, 1H,
J5′′,6′′ = 6.0 Hz, J5′′,3′′ = 2.4 Hz, H-5′′), 8.30 (d, 1H, J6′′,5′′ = 8.7 Hz, H-6′′), 8.15 (d, 1H, J4′ (3′,5′) = 9.0
Hz, H-4′), 7.56 (m, 8H, H-2, H-6, H-2′, H-6′, H-3, H-5, H-3′, H-5′); FAB-MS m/z 427 [M+H]+.
4'-Chloro-N'-(diphenylmethylene)-3''-nitrobenzenesulfonohydrazide (25)
Yield: 87%; m.p 168-170 °C; Rf: 0.36 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.76 (s, 1H, NH), 8.56 (d, 1H, J2′′,4′′ = 2.0 Hz, H-2′′), 8.19 (dd, 1H, J5′′,6′′ = 6.6 Hz, J5′′,2′′
=
2.0 Hz, H-5′′), 8.09 (d, 1H, J6′′,5′′ = 8.4 Hz, H-6′′), 7.54 (m, 3H, H-2, H-4, H-6), 7.43 (m, 4H, H-3,
13
H-5, H-3′, H-5′), 7.29 (m, 3H, H-2′, H-6′, H-4′); C-NMR: (100.0 MHz, DMSO-d6): C 156.6
(C=N), 147.1 (C-1′′), 138.5 (C-3′′), 136.8 (C-4′′), 132.9 (C-1′), 132.4 (C-1), 132.3 (C-4′), 130.1
(C-2, C-6), 129.6 (C-2′′), 128.9 (C-3′′), 128.8 (C-3′, C-5′), 128.4 (C-4), 127.5 (C-2, C-6), 125.2
(C-3, C-5); FAB-MS m/z 416 [M+H]+.
3'',5''-Dichloro-N'-(diphenylmethylene)-2-hydroxybenzenesulfonohydrazide (26)
Yield: 89%; m.p 185-187 °C; Rf: 0.37 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 11.10 (s, 1H, NH), 9.88 (s, 1H, OH),7.94 (d, 1H, J6′′,4′′ = 2.7Hz, H-6′′), 7.72 (d, 1H, J4′′,6′′
=
2.4 Hz, H-4′′), 7.59 (m, 2H, H-2, H-6), 7.37 (m, 4H, H-4, H-4′, H-2′, H-6′), 7.29 (m, 4H, H-3, H-
5, H-3′, H-5′); FAB-MS m/z 421 [M+H]+.
28

N'-(Diphenylmethylene)-4''-methoxybenzenesulfonohydrazide (27)
Yield: 87%; m.p 160-162 °C; Rf: 0.38 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.27 (s, 1H, NH), 7.86 (d, 1H, J4′ (3′,5′) = 8.7 Hz, H-4′), 7.52 (m, 2H, H-3, H-5), 7.47 (m,
3H, H-2, H-4, H-6), 7.37 (m, 4H, H-2′, H-6′, H-3′, H-5′), 7.29 (m, 2H, H-2′′, H-6′′), 7.15 (d, 2H,
J3′′,2′′/5′′,6′′ = 8.7 Hz, H-3′′, H-5′′); FAB-MS m/z 367 [M+H]+.
N'-(Diphenylmethylene)-2′′,4′′-dimethoxybenzenesulfonohydrazide (28)
Yield: 90%; m.p 180-182 °C; Rf: 0.39 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 7.58 (d, 2H, J2,3/6,5 = 8.4 Hz, H-2, H-6), 7.47 (d, 2H, J2′,3′/6′,5′ = 7.2 Hz, H-2′, H-6′), 7.42
(m, 3H, H-4, H-4′, H-6′′), 7.29 (m, 2H, H-3, H-5), 6.45 (d, 2H, J3′′,5′′/5′′,3′′ = 2.4 Hz, H-3′′, H-5′′),
6.39 (m, 2H, H-3′, H-5′), 3.73 (s, 3H, 2′′-OCH3), 3.70 (s, 3H, 4′′-OCH3); 13C-NMR: (100.0 MHz,
DMSO-d6): C 160.9 (C=N), 159.0 (C-2′′), 157.3 (C-4′′), 137.4 (C-1′′), 135.0 (C-1′′), 129.9 (C-1′),
129.4 (C-1), 128.9 (C-2, C-6), 128.8 (C-2′, C-6′), 128.7 (C-3, C-5 ), 128.3 (C-3′′), 128.1 (C-5′′),
128.0 (C-4), 10.2.8 (C-3′, C-5′), 98.7 (C-4′); FAB-MS m/z 397 [M+H]+.
N'-(Diphenylmethylene)naphthalene-2-sulfonohydrazide (29)
Yield: 88%; m.p 256-258 °C; Rf: 0.40 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.57 (s, 1H, NH), 8.58 (s, 1H, H-1′′),8.23 (d, 1H, J5′′,6′′ = 7.2Hz, H-5′′), 8.16 (d, 1H, J8′′,7′′
= 8.7 Hz, H-8′′), 8.06 (d, 1H, J4′′,3′′ = 8.7Hz, H-4′′), 7.96 (dd, 1H, J3′′,4′′ = 6.0 Hz, J3′′,1′′ = 2.0 Hz, H-
3′′), 7.74 (m, 3H, H-2, H-4, H-6), 7.35 (m, 2H, H-3, H-5), 7.35 (m, 2H, H-6′′, H-7′′), 7.33 (m, 5H,
13
H-2′, H-3′, H-4′, H-5′, H-6′); C-NMR: (75.0 MHz, DMSO-d6): C 159.0 (C=N), 154.7 (C-1′′),
137.4 (C-4a′′), 135.9 (C-2′′), 135.0 (C-2, C-6), 132.5 (C-8′′), 131.6 (C-8a), 129.9 (C-2′, C-6′),
129.3 (C-8a′′), 129.7 (C-3′, C-5′), 129.3 (C-4), 129.2 (C-3, C-5), 128.9 (C-1), 128.8 (C-1′), 128.3
(C-6′′), 128.1 (C-4′), 128.0 (C-5′′), 127.8 (C-3′′), 127.5 (C-4′′), 127.2 (C-4), 123.1 (C-7′′) ; EI-MS
m/z (% rel. abund.): 386 [M]+ (59.5), 321, (6.9), 195, (100.0), 166, (77.0), 128, (52.2), 92, (64.4).
5-(Dimethylamino)-N'-(diphenylmethylene)naphthalene-1-sulfonohydrazide (30)
Yield: 86%; m.p 250-252 °C; Rf: 0.39 (ethyl acetate/hexanes, 4:6); 1H-NMR (300 MHz, DMSO-
d6): H 10.84 (s, 1H, NH), 8.55 (t, 2H, J3′′ (2′′,4′′)/7′′ (6′′,8′′) = 9.6 Hz H-3′′, H-7′′), 8.24 (d, 1H, J2′′,3′′
=
8.7 Hz, H-4′′), 7.71, (d, 1H, J8′′,7′′ = 8.4 Hz, H-8′′), 7.65 (d, 1H, J2′′,3′′ = 6.0 Hz, H-2′′), 7.61 (d, 1H,
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