Novel Schiff bases derived from N-aryl maleimide derivatives as an effective antimicrobial agent: Theoretical and experimental approach

Article abstract of DOI:10.1016/j.bioorg.2020.104129

A set of new Schiff bases of N-aryl 3- and 4-substituted maleimides has been prepared via condensation of N-aryl 3- and 4- substituted maleimides with p-toluene sulfonyl hydrazide in acidic medium at room temperature. The structures of synthesized compounds were characterized by IR, ¹H NMR, ¹³C NMR, MS spectral data, and further confirmed by single-crystal x-ray crystallography for 5c. The computational study was carried out using Gaussian 09 software by using the B3LYP/6-311+G(d,p) basis set. Single-crystal study results showed much closeness with computational study results. These novel compounds were screened for their antimicrobial activity against two pathogenic bacteria such as Escherichia coli (ATCC 8739) and Staphylococcus aureus (ATCC25923) and two pathogenic fungi such as Aspergillus niger (MCIM10231) and Candida albicans (MTCC6275). The investigation of antimicrobial screening data showed that the most of tested compounds are moderate to good microbial inhibitors.

Full text of DOI:10.1016/j.bioorg.2020.104129

Journal Pre-proofs
Novel Schiff bases derived from N-aryl maleimide derivatives as an effective
antimicrobial agent: Theoretical and experimental approach
Arun M. Bhagare, Jayraj S. Aher, Manoj R. Gaware, Dnyaneshwar D.
Lokhande, Anant V. Kardel, Avinash D. Bholay, Akshay C. Dhayagude
PII:
S0045-2068(20)31426-7
DOI:
https://doi.org/10.1016/j.bioorg.2020.104129
YBIOO 104129
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To appear in:
Bioorganic Chemistry
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Revised Date:
Accepted Date:
14 May 2020
7 July 2020
18 July 2020
Please cite this article as: A.M. Bhagare, J.S. Aher, M.R. Gaware, D.D. Lokhande, A.V. Kardel, A.D. Bholay,
A.C. Dhayagude, Novel Schiff bases derived from N-aryl maleimide derivatives as an effective antimicrobial
agent: Theoretical and experimental approach, Bioorganic Chemistry (2020), doi: https://doi.org/10.1016/
j.bioorg.2020.104129
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Novel Schiff bases derived from N-aryl maleimide derivatives as an effective
antimicrobial agent: Theoretical and experimental approach
Arun M. Bhagare1, Jayraj S. Aher2*, Manoj R. Gaware3, Dnyaneshwar D. Lokhande3,
Anant V. Kardel4, Avinash D. Bholay5, Akshay C. Dhayagude1
1Department of Chemistry, MVP Samaj’s K.K. Wagh Arts, Science, and Commerce College,
Pimpalgaon (B.), Nashik-422 209, Maharashtra, India.
2Post Graduate Department of Chemistry, K.R.T. Arts, B.H. Commerce, and A.M. Science
College, Nashik-422002, Maharashtra, India.
3Department of Chemistry, MVP Samaj’s K.P.G. Arts, Science, and Commerce College, Igatpuri,
Nashik-422 403, Maharashtra, India.
4Department of Chemistry, MVP Samaj’s S.S.S.M. Arts, Science, and Commerce College,
Saikheda, Nashik-422 210, Maharashtra, India
5Post Graduate Department of Microbiology, K.R.T. Arts, B.H. Commerce, and A.M. Science
College, Nashik-422002, Maharashtra, India.
ABSTRACT:
A set of new Schiff bases of N-aryl 3- and 4-substituted maleimides has been prepared via
condensation of N-aryl 3- and 4- substituted maleimides with p-toluene sulfonyl hydrazide in
acidic medium at room temperature. The structures of synthesized compounds were
1
characterized by IR, H NMR, 13C NMR, MS spectral data, and further confirmed by single-
crystal x-ray crystallography for 5c. The computational study was carried out using Gaussian 09
software by using the B3LYP/6-311+G(d,p) basis set. Single-crystal study results showed much
closeness with computational study results. These novel compounds were screened for their
antimicrobial activity against two pathogenic bacteria such as Escherichia coli (ATCC 8739) and
Staphylococcus aureus (ATCC25923) and two pathogenic fungi such as Aspergillus niger
(MCIM10231) and Candida albicans (MTCC6275). The investigation of antimicrobial screening
data showed that the most of tested compounds are moderate to good microbial inhibitors.
Keywords: Schiff base, N-aryl 3- and 4-substituted maleimide, p-toluene sulfonyl hydrazide,
antibacterial, antifungal activity
1. Introduction
This is the era in which various invading microorganisms threaten to mankind by their adverse
effects. Antimicrobial resistance by this microbial organism is a new challenge for the
pharmaceutical industry and clinicians. The Multi-Drug-Resistance (MDR) bacteria and fungi
are some of the important factors for the failure of chemotherapy in the number of infectious
disease1,2. Heterocyclic compounds are widely used in the pharmaceutical industry due to their
physicochemical properties, selective affinity towards biomolecules, and constructive properties

as therapeutics agents. Moreover, synthetic drugs like diazepam, chlorpromazine, metronidazole,
antipyrine, isoniazid, etc. are well-known heterocycles3,4.They are also an efficient antifungal5,
anti HIVactivity6, antimalarial7, antimycobacterial8, antibacterial9, analgesic, and anti-
inflammatory agent10. Therefore, nowadays synthesis of new heterocyclic compounds and their
variety of derivatives became a thrust area for the organic chemist. Hence, every year lot of
efforts were made to synthesize new organic heterocyclic compounds.
Recently many researchers reported that maleimide derivatives can be used as selective
inhibitors for monoglyceridelipase11, Cdc25B12, GSK-3α13, Bfl-114and DNMT-115. Moreover,
Maleimides are one of the promising group of heterocyclic compounds containing -CO-N(R)-
CO-chain. Due to their hydrophobic and neutral nature, it can easily penetrate through biological
membrane16. Hence widely applied for various biological applications like antibacterial17,
antimicrobial18, antiprotozoal19, analgesics20, antitiangiogenic21, antistressagents22, cytotoxic,
DNA binding and apoptotic inducing activity23. Maleimides are the emerging class of
heterocyclic compounds and showed wide biological applications. Therefore many researches
are motivated to synthesize maleimide derivatives.
S. Zacchino and co-workers reported the antimicrobial activity of N- substituted maleimide
moiety against Candida spp. They observed that the maleimide framework is playing a vital role
in enhancing the biological activity of thesecompounds24.N. Salewskaet.al was reported the
structure-dependent antifungal activity of N- substituted maleimides25. N.S. Patil and group were
synthesized α-hydroxy phosphonates clubbed derivative of N-aryl maleimides and investigated
their antimicrobial activity towards Bacillus subtilis, Escherichia coli bacteria and Candida
Albicans, Candida tropicalis, Aspergillusniger and Aspergillus clavatus fungi26.
The compounds containing azomethine (CH=N-) groups are generally known as Schiff bases.
They are synthesized by condensation of carbonyl compounds with primary amines.Such
compounds possess good biological properties due to the presence of the CH=N group. They are
good intermediates in enzyme-catalyzed reactions of amino or carbonyl group substrates27. They
also possess good excellent biological activities such as antibacterial28, antifungal29,
antimalarial30, antitumor31, antiviral32, antioxidants33, analgesic, and anti-inflammatory34.
However, clubbing of both maleimide derivative and Schiff base may enhance their biological
property due to the synergistic effect.

In the present study, we have reported the facile synthesis of new Schiff base derivatives of N-
aryl 3- and4- substituted maleimides by condensation with p-toluene sulfonyl hydrazide. Most of
0
the research group reported synthesis at 50-70 C 35,36. But here we have performed synthesis at
room temperature. Moreover, synthesized derivatives have good yield and crystallinity. The
synthesized derivative was characterized by different analytical techniques such as 1H NMR, 13C
NMR, IR, mass spectroscopy, and single-crystal x-ray diffractometer. The experimental results
were supported by density functional theory (DFT) calculation with the help of the Gaussian 09
software package. The antibacterial activity of synthesized derivatives was evaluated against
both gram-positive and gram-negative bacteria viz Escherichia coli (E. coli), Staphylococcus
aureus (S. aureus). The antifungal activity of the synthesized derivative was also evaluated
against fungi viz. Aspergillus Niger (A. niger) and Candida albicans (C. albicans). The results
showed enhanced activity than standard Fluconazole. Moreover, we observed those substituents
are playing a major role in biological activity. Thus present work may provide a new direction to
synthesize modified derivative of maleimides. Furthermore, it can be applied as a promising
candidate for various biological applications.
R
O
O
O
S
S
N
R
Ar
Ar
H
N
R
N
N
R1
O
R3
R2
O
O
R1
Antimicrobial18
GSK-3 inhibitors23
Antifungal13
R2
R3
O
O
H
N
O
R1
R2
S
N
R1
N
O
N
N
R
R4
O
O
Analgesic and Anti Infammatory34
Synthesized Compounds (5a-l)
Figure 1. Chemical structures of some biologically active maleimide derivatives and Schiff base
scaffold

O
R3
R2
O
R3
R2
O
OHC
R4
R3
R2
ii
Br
Br
i
N
R1
N
R1
N
R1
R4
O
O
(2a-l)
(3a-l)
O
(1)
O
S
a) R1 = R2 = R3 = H; R4 = Piperidyl
b) R2 = R3 = H; R1= Cl; R4 = Piperidyl
c) R2 = R3 = H; R1= Br; R4 = Piperidyl
iii
NH
NH2
O
(4)
d) R2 = R3 = H; R1= F; R4 = Piperidyl
e) R2 = R3 = H; R1= CH3; R4 = Piperidyl
R2
R3
O
H
O
f) R2 = R3 = H; R1= NO2; R4 = Piperidyl
g) R1 = R2 = R3 = H; R4 = N(Me)2
N
S
N
R1
N
h) R2 = R3 = H; R1= Cl; R4 = N(Me)2
O
i) R2 = R3 = H; R1= Br; R4 = N(Me)2
j) R2 = R3 = H; R1= F; R4 = N(Me)2
k) R2 = R3 = H; R1= CH3; R4 = N(Me)2
R4
(5a-l)
O
l) R1 = R3 = H; R2= NO2; R4 = N(Me)2
Scheme 1: Reagent and conditions; (i) N,N-dimethylamine / Piperidine, DMF,0-10 oC,30 min.; (ii) DMF,
POCl3,0-5 oC, 30 min.; (iii) Ethanol, Acetic acid,30 min.
Table 1: Melting point, practical yield and approximate IR frequencies of the compounds (5a-l)
Entry
R1
R2
R3
R4
M.P.(oC)
Yield
(%)
92
94
92
90
95
90
94
93
94
91
95
ʋ/cm-1
C=N N-H
C=O
5a
5b
5c
5d
5e
5f
5g
5h
5i
H
Cl
H
H
H
H
H
H
H
H
H
H
H
NO2
H
Piperidyl 170-172
Piperidyl 168-170
Piperidyl 161-163
Piperidyl 177-179
Piperidyl 163-165
Piperidyl 191-193
N(Me)2
N(Me)2
N(Me)2
N(Me)2
N(Me)2
N(Me)2
1752, 1686
1752, 1692
1757, 1689
1754, 1701
1748, 1688
1756, 1698
1761, 1690
1755, 1701
1753, 1694
1744, 1688
1753, 1696
1759, 1701
1602 3432
1607 3465
1602 3439
1612 3193
1601 3435
1605 3455
1609 3387
1612 3472
1605 3421
1604 3265
1607 3415
1612 3460
H
H
H
H
H
H
H
H
H
H
H
Br
F
CH3
NO2
H
Cl
Br
F
177-180
202-204
194-196
192-194
178-180
201-203
5j
5k
5l
CH3
H
89

2. Result and Discussion
2.1 Spectroscopic analysis
IR spectrum of the synthesized compounds (5a-l) showed the stretching frequencies for the
carbonyl group at 1743-1761 cm-1 and 1687-1701 cm-1. The IR spectra show a strong band at
1602-1612 cm-1 which is attributed to the presence of azomethine (C=N) group. IR spectrum also
1
shows N-H stretching frequency at 3387-3439 cm-1. The H NMR spectrum (DMSOd6) of
synthesized compound(5a-f) showed broad singlet at the range δ 1.53-1.54 ppm (J= 4 Hz) for
four protons of two –CH2- groups, broad singlet at the range δ 1.58-1.59 ppm (J= 4-5 Hz) for
two protons of one –CH2 group and a broad singlet at the range δ 3.68-3.69 ppm for four protons
of two –CH2 group of piperidine nucleus whereas for compounds (5g-l) showed broad singlet at
1
the range δ 3.21-3.24 ppm for six protons of –N(CH3)2 group. The H NMR peaks singlet at
7.809-7.842 ppm attributed the presence of -CH=N- group. The 1HNMR spectrum showed peaks
singlet at 11.252-11.281 ppm are due to the presence of –NH- proton whereas singlet at 2.382-
2.389 ppm is due to –CH3 group of toluene side chain. The peaks in the range 7.275-7.833 ppm
are attributed to the aromatic protons.
The 13C NMR spectrum of synthesized compounds (5a-e) showed peaks in the range of 22.94-
23.54 ppm for one carbon, 26.25-26.85 ppm for two carbons and 51.17-51.84 ppm for two
carbon of piperidine framework while for compounds (5g-l) showed characteristic peaks in the
range 43.10-43.92 ppm for six carbons of dimethylamine group. The 13C NMR spectrum showed
peaks in the range 161-169 ppm for carbonyl (C=O) groups and 144.31-145.98 ppm for the –
C=N- group. The peaks at the range 93-94 ppm attributed to the presence of -CH= group
whereas the peaks at the range of 20-26 ppm showed the presence of –CH3 group of Toluene
nucleus.
2.2 Crystal Structure of 5c
The molecular structure of the compound 5c with the numbering scheme is given in Figure (2).
The technical data collection report and some selected parameters of the refinement of
compound 5c are tabulated in Table 2 and some selected bond angles and the bond length also
shown in Table 3. The compound 5c possesses a triclinic crystal system with a P-1 space group.

The molecular density of compound 5c was found to be 1.515 mg m-3and which is in agreement
with the theoretical expected value 1.53±0.1 mg m-3. The crystal structure of 5c shows C8-N2
distance 1.268 Å for the C=N double bond. The Schiff base molecule adopted E-configuration in
connection with the C=N bond with N1-N2-C8 bond angle 117.8o and N2-C8-C9 bond angle
124.3o. Crystal packing visualization of compound 5c along different axis is shown in Figure 3
and Figure 4.
Figure 2: ORTEP diagram of compound 5c

Figure 3: Crystal packing of the 5c viewed along the axis
Figure 4: Crystal packing of compound 5c along the A-axis

Table 2: Crystal data and structure refinement for compound 5c
Empirical formula
Formula weight
Temperature
C23 H23 Br N4 O4 S
531.42
273(2) K
Wavelength
71.073 pm
Crystal system
Space group
Triclinic
P -1
Unit cell dimensions
a = 785.57(6) pm
b = 1092.13(6) pm
c = 1402.32(8) pm
α= 86.521(2)°.
β= 88.989(2)°.
γ = 75.993(2)°.
3
Volume
1.16518(13) nm
2
Z
3
Density (calculated)
Absorption coefficient
F(000)
1.515 Mg/m
-1
1.891 mm
544
3
Crystal size
0.350 x 0.250 x 0.122 mm
2.911 to 28.422°.
Theta range for data collection
Index ranges
-10<=h<=10, -14<=k<=14, -18<=l<=18
Reflections collected
Independent reflections
Completeness to theta = 25.242°
Refinement method
Data / restraints / parameters
15418
5833 [R(int) = 0.0303]
99.3 %
2
Full-matrix least-squares on F
5833 / 0 / 300
2
Goodness-of-fit on F
1.024
Final R indices [I>2sigma(I)]
R indices (all data)
R1 = 0.0564, wR2 = 0.1282
R1 = 0.1020, wR2 = 0.1505
0.018(2)
Extinction coefficient
-3
1.088 and -1.338 e.Å
Largest diff. peak and hole

Table 3: Comparison of bond lengths (Å) for compound 5c
Bond Lengths(Å)
B3LYP/6-311+G(d,p) Experimental Bond Lengths(Å) B3LYP/6-311+G(d,p) Experimental
C1-C2
1.3966
1.3966
1.425
1.3860
1.3780
1.4270
1.3860
0.9300
1.3650
0.9300
1.3790
1.8950
1.3770
0.9300
0.9300
1.4070
1.3850
1.4620
1.2130
1.5140
1.2020
1.3800
1.4460
1.3390
1.5070
0.9700
1.4630
0.9700
1.5070
0.9700
1.4770
0.9700
1.5080
C18-H23
C18-H49
C20-C22
C20-H24
C20-H47
C22-H25
C22-H48
C26-H27
C26-N38
C28-C29
C28-C30
C28-S51
C29-C31
C29-H32
C30-C33
C30-H34
C31-C35
C31-H36
C33-C35
C33-H37
C35-C52
N38-N39
N39-H40
N39-S51
O41-S51
O50-S51
C52-H53
C52-H54
C52-H55
1.0955
1.0946
1.5329
1.095
0.9700
0.9700
1.5080
0.9700
0.9700
0.9700
0.9700
0.9300
1.2680
1.3680
1.3720
1.7500
1.3780
0.9300
1.4030
0.9300
1.3420
0.9300
1.3760
0.9300
1.5290
1.4040
0.8600
1.6480
1.4180
1.4300
0.9600
0.9600
0.9600
C1-C6
C1-N11
C2-C3
1.3916
1.0811
1.3909
1.0821
1.3907
1.9178
1.3919
1.0821
1.0808
1.4382
1.3834
1.4805
1.2017
1.5262
1.2118
1.3754
1.4514
1.3552
1.5347
1.0881
1.4688
1.0999
1.529
C2-H7
1.0942
1.0936
1.0978
1.0961
1.2815
1.3914
1.3948
1.7938
1.3935
1.0829
1.389
C3-C4
C3-H8
C4-C5
C4-Br56
C5-C6
C5-H9
C6-H10
N11-C12
N11-C13
C12-C14
C12-O44
C13-C15
C13-O43
C14-C15
C14-C26
C15-N42
C16-C18
C16-H19
C16-N42
C16-H45
C17-C20
C17-H21
C17-N42
C17-H46
C18-C22
1.083
1.3984
1.0847
1.4027
1.0849
1.5081
1.3726
1.0254
1.7425
1.4619
1.4519
1.0926
1.0952
1.0916
1.1008
1.4766
1.0843
1.532

Table 4: Comparison between theoretical and experimental bond angles of compound 5c
Bond angles(o)
C2-C1-C6
B3LYP/6-311+G(d,p) Experimental Bond angles(o)
B3LYP/6-311+G(d,p)
Experimental
109.2
109.2
109.2
109.2
107.9
110.0
109.7
109.7
109.7
109.7
108.2
117.8
124.3
117.8
119.9
120.7
119.4
118.6
120.7
120.7
120.0
120.0
120.0
121.1
119.4
119.4
121.9
119.1
119.1
118.5
120.0
120.8
119.2
119.4
120.3
120.3
120.0
120.0
120.0
120.9
118.7
120.4
119.4
120.3
120.3
120.3
119.8
119.8
123.2
126.8
109.9
108.0
128.2
123.8
106.6
125.4
127.9
108.1
116.8
134.5
120.0
120.2
119.9
120.2
120.1
119.7
119.4
120.1
120.5
121.0
119.5
119.5
119.4
120.5
120.0
120.1
120.1
119.8
124.6
125.1
110.4
106.8
123.5
129.7
106.4
126.3
127.2
108.4
122.8
128.4
C17-C20-H24
C17-C20-H47
C22-C20-H24
C22-C20-H47
H24-C20-H47
C18-C22-C20
C18-C22-H25
C18-C22-H48
C20-C22-H25
C20-C22-H48
H25-C22-H48
C14-C26-H27
C14-C26-N38
H27-C26-N38
C29-C28-C30
C29-C28-S51
C30-C28-S51
C28-C29-C31
C28-C29-H32
C31-C29-H32
C28-C30-C33
C28-C30-H34
C33-C30-H34
C29-C31-C35
C29-C31-H36
H35-C31-H36
C30-C33-C35
C30-C33-H37
H35-C33-H37
C31-C35-C33
108.8
108.7
110.2
110.8
107.2
110.4
110.3
109.4
110.6
109.5
106.6
116.9
122.7
120.4
121.2
119.5
119.3
119.0
119.8
121.2
118.9
120.1
121.0
121.6
119.2
119.6
121.2
119.3
119.5
118.5
C2-C1-N11
C6-C1-N11
C1-C2-C3
C1-C2-H7
C3-C2-H7
C2-C3-C4
C2-C3-H8
C4-C3-H8
C3-C4-C5
C3-C4-Br56
C5-C4- Br 56
C4-C5-C6
C4-C5-H9
C6-C5-H9
C1-C6-C5
C1-C6-H10
C5-C6-H10
C1-N11-C12
C1-N11-C13
C12-N11-C13
N11-C12-C14
N11-C12-O44
C14-C12-O44
N11-C13-C15
N11-C13-O43
C15-C13-O43
C12-C14-C15
C12-C14-C26
C15-C14-C26

107.3
120.0
132.6
109.2
112.0
109.7
109.2
107.9
109.2
109.6
110.4
109.6
109.6
108.1
109.6
111.5
109.3
109.3
109.3
109.5
108.0
112.3
119.3
122.2
115.2
123.4
113.1
123.4
123.6
122.4
113.5
105.7
108.5
109.6
108.6
103.8
119.7
109.5
109.5
109.5
109.5
109.5
109.5
C13-C15-C14
C13-C15-N42
C14-C15-N42
C18-C16-H19
C18-C16-N42
C18-C16-H45
H19-C16-N42
H19-C16-H45
N42-C16-H45
C20-C17-H21
C20-C17-N42
C20-C17-H46
H21-C17-N42
H21-C17-H46
N42-C17-H46
C16-C18-C22
C16-C18-H23
C16-C18-H49
C22-C18-H23
C22-C18-H49
H23-C18-H49
C17-C20-C22
108.0
120.5
131.2
109.7
110.6
109.8
108.8
108.0
109.9
109.6
111.5
110.9
107.3
108.4
109.1
111.9
108.6
108.4
109.8
110.9
107.1
111.0
C31-C35-C52
C33-C35-C52
C26-N38-N39
N38-N39-H40
N38-N39-S51
H40-N39-S51
C15-N42-C16
C15-N42-C17
C16-N42-C17
C28-S51-N39
C28-S51-O41
C28-S51-O50
N39-S51-O41
N39-S51-O50
O41-S51-O50
C35-C52-H53
C35-C52-H54
C35-C52-H55
H53-C52-H54
H53-C52-H55
H54-C52-H55
121.0
120.5
116.6
115.4
113.7
109.0
122.3
122.8
114.1
98.2
108.0
110.1
109.9
106.8
121.4
111.4
110.6
111.5
107.3
108.3
107.6
2.3 Computational Study
The computational study of compound 5c was carried out by using the Gaussian 09 software by
using the B3LYP/6-311+G(d,p) basis set. The optimized structure of compound 5c is shown in
Figure 5. The optimized parameters obtained by the theoretical study were compared with the
experimental parameters of single-crystal study results. The values of the parameters like bond
angle, bond length are much closer to each other, and as shown in Table 3 and Table 4. Thus
theoretical results corroborate very well with the experimental results.

Figure 5: Optimized structure of the compound 5c
Here, the charge distribution has been characterized by Molecular electrostatic surface potentials
(MESP) mapped topography as illustrated in Figure. 6.
Figure 6: MESP plot by the DFT/B3LYP method with the 6-311+G(d,p) as a basis set

The effective electron-rich regions (red) are observed in the vicinity of the lone pair residing on
oxygen atom centers. Whereas, more electron-deficient region was identified near the bromine
groups than the aromatic hydrogens. The net Mulliken charges on each atom were obtained from
population analysis and given in Table S3, which further corroborates these inferences. The
gradual variation of electron-rich regions with varying functional groups on 5c comprising
bromine atom is evident from Figure. 6. Moreover, the intense aromatic C=O stretching of the
imide group was assigned to a band near ~1698 cm−1 which is consistent with experimental
values. The intense vibration of aromatic C-N is observed 1347 cm−1. Furthermore to
substantiate the stability and reactivity of the proposed molecule frontier molecular orbital
analysis was performed. Figure 7 represents the HOMO-LUMO plots of compound 5c. The
frontier orbital’s LUMO resides near the central five-membered ring consisting of imide
functionality. On the other hand frontier orbital’s HOMO is mostly located over bromobenzene
and piperidine ring of the synthesized compound 5c. The thermodynamic parameters (Table S1)
and Cartesian coordinates (Table S5) are given separately in supporting the information table.

ELUMO = -0.09699 a.u.
∆E = 0.12531 a. u.
.
EHOMO = -0.22230 a.u.
Figure 7: The HOMO and LUMO plots by the DFT/B3LYP method with 6-311+G(d,p)

2.4 Antimicrobial Activity/ screening
Maleimide derivatives are showing efficient antimicrobial activity due to their hydrophobic and
neutral nature. Moreover, Schiff bases are also known as excellent antimicrobial agents. To study
the synergic effect due to clubbing of N-aryl 3- and 4-substituted maleimides with Schiff base,
the antimicrobial activity of the synthesized compounds was examined against bacteria (E. coli
S. aureus) and fungi ( A. niger, C. albicans). To choose a suitable concentration for antimicrobial
study, concentration-dependent activity was examined against pathogenic bacteria and fungi.
Initially, antimicrobial activity was checked for the one of the synthesized derivative (5c) at
different concentrations viz. 10, 20, and 30 µg mL-1. We noticed that at 20 and 30 µg mL-1
concentration results are almost similar whereas, at 10 µg mL-1 concentration zone of inhibition
was relatively less than other concentrations (20,30 µg mL-1). Therefore, the antimicrobial
activity of other derivatives was examined at 20 µg mL-1 concentration.
Table 5: Antimicrobial screening of synthesized compounds 5a-l (zone diameter of growth inhibition in
mm)
Entry
Escherichia coli Staphylococcus aureus Aspergillusniger Candida albicans
ATCC 8739
15 ±0.8
15 ±0.5
14 ±0.3
14 ±0.2
13 ±0.5
10 ±0.4
12 ±0.7
13 ±0.3
13 ±0.5
12 ±0.8
11 ±0.4
11 ±0.5
8 ±0.4
ATCC25923
10 ±0.2
12 ±0.5
11 ±0.3
12 ±0.7
11 ±0.8
10 ±0.2
13 ±0.8
14 ±0.4
15 ±0.3
12 ±0.6
11 ±0.2
12 ±0.7
8 ±0.4
MCIM10231
22 ±0.8
17 ±0.9
16 ±0.3
26 ±0.5
24 ±0.6
16 ±0.2
16 ±0.4
17 ±0.2
18 ±0.7
15 ±0.5
14 ±0.8
16 ±1.1
9 ±0.3
MTCC6275
15 ±0.8
14 ±0.2
14 ±0.4
15 ±1.2
14 ±0.6
15 ±0.4
13 ±0.2
13 ±0.5
14 ±0.7
12 ±0.3
11 ±0.4
11 ±0.6
9 ±0.6
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
DMSO
Gentamicin
(20µg mL-1)
Fluconazole
(20µg mL-1)
22 ±0.3
23 ±0.5
-
-
-
-
24 ±0.2
22 ±0.4
The antibacterial activity was examined against the gram-negative E. coli while gram-positive S.
aureus and observed results were summarised in Table 5. The inhibition zone diameter of

dimethylamine substituted derivatives (5g-l) against E. coli was found to be 12±0.7, 13±0.3,
13±0.5, 12 ±0.8,11 ±0.4 and 11 ±0.5 mm respectively whereas, for piperidine substituted
derivative (5a-f), it was 14 ±0.3, 15 ±0.5, 14 ±0.2, 15 ±0.8, 13 ±0.4 and 10 ±0.4 mm
respectively. The diameter of inhibition zone against S. aureus bacteria in dimethylamine
substituted derivatives (5g-l) was observed to be 13 ±0.8, 14 ±0.4, 15 ±0.3, 12 ±0.6, 11 ±0.2 and
12 ±0.7 mm and for piperidine substituted compounds(5a-f), it was 11 ±0.3, 12 ±0.5, 12 ±0.7, 10
±0.2, 11 ±0.8 and 10±0.2 mm respectively. The bacterial study indicate that the dimethylamine
substituted compounds (5g-l) showed higher activity against gram-positive bacteria (S.aureus)
whereas, piperidine substituted compounds (5a-e) showed better activity against gram-negative
bacteria (E. coli).
The antifungal activity of synthesized derivatives (5a-l) was also tested against A. niger and C.
albicans fungi and the observed results were summarized in table 5.The inhibition zone diameter
due to piperidine substituted derivatives (5a-f) against A. niger was found to be 16 ±0.3, 17 ±0.9,
26 ±0.5, 22 ±0.8, 24 ±0.6 and 16 ±0.3 mm respectively. Similarly, the inhibition zone against C.
albicans was observed 14 ±0.4, 14 ±0.2, 15±0.2, 15±0.6, 14 ±0.6, and 15±0.4 mm respectively.
The inhibition zone diameter of dimethylamine substituted compounds (5g-l) towards A. niger
was observed to be 16 ±0.4, 17 ±0.2, 18 ±0.7, 15±0.5, 14±0.8, 16±0.1 mm respectively. Against
C. albicans, it was observed to be 13 ±0.2, 13±0.5, 14 ±0.7, 12 ±0.4 and 11 ±0.6 mm
respectively.
For antimicrobial study, the solution of the synthesized derivative was prepared in DMSO.
Therefore the activity of the DMSO was also investigated as a control. The control sample
showed 8 ±0.4 mm inhibition zones against E. coli and S. aureus bacteria and 9 ±0.6 mm against
A. niger and C. albicans fungi.
From our present antimicrobial study, it was noticed that the substituent playing an important
role. The antifungal activity of dimethylamine substituted derivatives 5g, 5h, 5i was found to be
16 ±0.7, 15 ±0.5, and 14±0.8 mm however for piperidine substituted derivatives 5a, 5d, 5e it is
observed 22 ±0.8, 26 ±0.8 and 24 ±0.6 mm respectively. The improvement in the antifungal
activity may be due to piperidine substituent or better interaction of the fungi with piperidine
substituted derivatives. A similar observation was also reported by N.Salewska and co-workers24.
They were synthesized several N-substituted maleimides and tested for antimicrobial and

cytostatic activity. They were observed that neutral maleimides displayed relatively strong
antifungal effects and their antibacterial activity was structure-dependent. The slightly enhanced
antibacterial activity of piperidine substituted derivatives was also observed against E.coli. From
antimicrobial study, it was noticed that substituents have more effect on antifungal activity than
antibacterial activity.
To compare our result with the standard antibacterial agent gentamicin, its antibacterial activity
was also recorded under identical conditions. It was found to be 22 ±0.3 and 23±0.5 mm against
E. coli and S. aureus bacteria respectively. Similarly, antifungal activity of fluconazole was also
recorded against A. niger and C. albicans and it was observed 24 ±0.2 and 24 ±0.4 mm
respectively. The most important outcome from our present study is that the some of the
piperidine substituted derivatives 5a, 5d, 5e showed equal / greater antifungal activity than the
standard fluconazole. The observed difference in antibacterial activity may be due to the
different interaction of the bacteria/fungi with the standard compounds or the different molecular
composition of the gram-positive and gram-negative bacteria. A detailed study will require
elucidating the mechanism of antibacterial activity. From our data, it is clear that synthesized
derivatives can be used as an antimicrobial agent for both gram-positive and gram-negative
bacteria and fungi.
3. Conclusions
A series of new Schiff bases of N-aryl 3- and 4- substituted maleimides (5a-l) were successfully
synthesized by condensation of N-aryl 3- and 4- substituted maleimides with p-toluene sulphonyl
hydrazide in ethanol in the acidic medium at room temperature. The novelty of this work is the
synthesis was performed at mild condition i.e. room temperature than other reports. The
synthesized compounds have excellent practical yield and good crystallinity. The purified
compound was characterized by different spectroscopic techniques.The single-crystal X-ray
1
diffraction study reveals a triclinic crystal structure with a P-1 space group. The H NMR and
13C NMR results confirm the formation of the proposed compound. Moreover, they also support
the high purity of the compound. The molecular weight obtained from mass spectroscopy was in
very good agreement with those obtained from theoretical (calculated). The synthesized
compound showed good antibacterial activity against Escherichia coli and Staphylococcus
aureus bacteria. Moreover, they also showed efficient antifungal activity towards Aspergillus

niger and Candida albicans fungi. From antimicrobial evaluation, it was observed that
substituent has some effect on activity. It worth mentioning here that some of the derivative(5a,
5d, 5e) showed equal / greater antifungal activity than those of standard Fluconazole.We assume
that due to good yield, high purity and efficient antimicrobial activity of synthesized compounds
can be used for different biological applications. Furthermore, our study may provide a new
direction to chemists (researchers) for the synthesis of newly substituted maleimide derivatives.
4. Experimental Section
4.1. Materials
All the reagents and chemicals used for the synthesis were analytical grade (AR). P-toluene
sulfonyl hydrazide was purchased from Spectrochem, India. N, N-Dimethyl formamide (DMF),
phosphorus oxychloride, acetic acid, maleic anhydride and dimethyl sulphoxide (DMSO) were
purchased from Merck chemicals India. Different substituted aniline was purchased from SD-
fine chemicals, India. Thin-layer chromatography (TLC) plate of 0.2 mm thin silica layer was
purchased from Merck Chemicals, India. All the solvents were used after purification by
standard literature procedures. Bacterial and fungal cultures were purchased from NCIM, NCL
Pune India. Muller Hinton agar and Czapek Dox agar were purchased from Hi-media, India.
4.2. Synthetic method
General procedure for the synthesis of compound (1)
The solutions of various substituted anilines (0.01 mol) were prepared in 10 mL acetic acid.
Then maleic anhydride (0.01 mol) was added with constant magnetic stirring. To this reaction
mixture bromine (0.011mol) was added dropwise. Further in the same pot sulfuric acid (0.025
mmol) was added and the reaction mixture was stirred for 30-45 minutes. Then the reaction
mixture was cooled at room temperature and poured onto ice-cold water. The solid was
separated, filtered, and washed with sodium bicarbonate followed by water and recrystallized by
aqeous1:1 ethyl alcohol37.

General procedure for the synthesis of compound (2a-l)
The solution of compound I (0.01 mol) was prepared in 10 mL purified DMF. In this solution
o
0.031 mol dimethylamine / piperidine was added dropwise at 0-10 C with constant magnetic
stirring. After completion of the reaction, the reaction mixture was poured into cold water. The
bright yellow solid appeared, then filtered and recrystallized by using ethanol26.
General procedure for the synthesis of compound (3a-l)
The compound 2a-l (0.01 mol) was dissolved in 2 mL DMF. Vilsmeier Haack adduct (DMF
o
(0.012 mol) and POCl3 (0.05 mol)) were added slowly dropwise at 0 C, then the reaction
o
mixture was stirred for 30 minutes at 0-5 C. The reaction mixture was poured into crushed ice
and neutralized by 10% NaHCO3. The yellow compound was obtained, which was filtered and
washed with cold water. The product was purified by recrystallization by using ethanol26.
General procedure for the synthesis of compound (5a-l)
5 mL ethanolic solution of synthesized compound 3a-l (0.01 mol) was prepared with the addition
of a small amount of acetic acid (0.005 mol). To the above solution 0.01 mol p-toluene, sulfonyl
hydrazide (4) was added slowly under constant stirring at room temperature. The initial
yellowish color of solution changes to transparent reddish. After ~20 minutes orange-red crystals
were obtained. The product was separated by filtration at the suction pump using Buchner
funnel. To remove impurity, the product was washed with purified cold ethanol. The shiny
orange crystals of compound 5a-l were purified by recrystallization using ethanol. The reaction
was monitored by TLC method using solvent n-hexane/ethyl acetate (1:1). The synthetic
procedure is schematically represented in Scheme 1.
4.3. Characterization
Synthesized compounds were purified and characterized by different analytical techniques.
Chemical composition information was obtained by recording the Infra-red (IR) spectrum on
Shimadzu Fourier transform infrared spectrophotometer. Nuclear magnetic resonance (NMR)
1
13
spectra were recorded for purification and structural information. The H-NMR and C-NMR
spectra were recorded on 500 MHz NMR spectrometer (BRUKER ADVANCE NEO, SAIF,
Panjab University, India). The values of chemical shifts (δ) were given in ppm relative to

tetramethylsilane (TMS) as standard. The multiplicities are abbreviated as s (singlet), brs (broad
singlet), d (doublet), t (triplet), q (quartet), or m (multiplet). Molecular weight and possible
fragmentation was estimated from Mass spectra (MS). MS was recorded on a mass spectrometer
(WATERS, Q-TOF MICROMASS (LC-MS)), SAIF, Punjab University, India). The crystal
structure was determined by Single Crystal- X-Ray Diffractometer (SCXRD), (Bruker, D8
VENTURE, S. P. P. U, Pune).
N'-{(E)-[4-(piperidin-1-yl)-2, 5-dioxo-1-phenyl-2,5-dihydro-1H-pyrrol-3-yl]methylidene}-4-
methylbenzene-1-sulfonohydrazide (5a)
1H NMR(500 MHz.DMSOd6) (δ-ppm) 1.53(br,4H, 2xCH2),1.59(br,2H, J=4.6Hz,CH2), 2.36
(s,3H, CH3), 3.68 (br,4H, 2xCH2), 7.26-7.43(m,4H, Ar-H), 7.64-7.81(m,5H, Ar-H), 7.82(s,1H,
CH=N), 11.29(s,1H,NH),13C NMR (500MHz, DMSOd6) 20.86 (CH3), 22.95 (1C,CH2),
26.26(2C’s, 2xCH2), 51.23(2C’s,2xCH2), 94.18(CH=), 120.21(Ar-C), 126.87(Ar-C), 127.21
(J=170Hz, (Ar-C)), 128.53(Ar-C),128.73 (J=100Hz, Ar-C),129.51(Ar-C),131.98(Ar-
C),131.51(Ar-C),136.16(Ar-C),139.73(Ar-C),139.84(J=55Hz,Ar-C),143.22(Ar-C),144.40(=C-
N),164.41()C=O), 168.76 (C=O), MS m/z : 452(M+),453 (M+1).
N'-{(E)-[1-(4-chlorophenyl)-4-(piperidin-1-yl)-2,5-dioxo-2,5-dihydro-1H-pyrrol-3-
yl]methylidene}-4-methylbenzene-1-sulfonohydrazide (5b)
1H NMR(500 MHz. DMSOd6) (δ-ppm) 1.53(br,4H, J=4Hz,2xCH2),1.59(br,2H, J=4.5Hz,CH2),
2.38(s,3H, CH3), 3.68(br,4H, 2xCH2), 7.32-7.35(m,2H, Ar-H), 7.42(d,2H,J=8Hz,Ar-H), 7.51-
7.53(m,2H,Ar-H)7.71(t,2H,J=6.5Hz,Ar-H), 7.82(s,1H, CH=N), 11.28(s,1H,NH),13C NMR
(500MHz, DMSOd6) 20.87(CH3), 22.95 (1C,CH2), 26.27(2C’s, 2xCH2), 51.50(2C’s,2xCH2),
94.21(CH=), 127.22(Ar-C),128.51(J=65Hz,Ar-C), 129.51(Ar-C), 130.56(Ar-C), 131.78(Ar-C),
136.19(Ar-C), 139.75(Ar-C), 143.22(Ar-C), 144.39(=C-N),164.48(C=O), 168.81(C=O), MS m/z
: 487(M+),488(M+1).
N'-{(E)-[1-(4-bromophenyl)-4-(piperidin-1-yl)-2,5-dioxo-2,5-dihydro-1H-pyrrol-3-
yl]methylidene}-4-methylbenzene-1-sulfonohydrazide (5c)
1H NMR(500 MHz. DMSOd6) (δ-ppm) 1.54(br,4H, J=4Hz,2xCH2), 1.59(br,2H, J=5Hz,CH2),
2.38(s,3H, CH3), 3.69(br,4H, 2xCH2), 7.29(t,2H,J=2Hz,Ar-H), 7.43(d,2H,J=8Hz, Ar-H),

7.66(t,2H,J=1.5Hz,Ar-H),7.71(d,2H,J=8.5Hz,Ar-H), 7.82(s,1H, CH=N), 11.28(s,1H,NH), 13C
NMR (500MHz, DMSOd6) 21.45(CH3), 23.54 (1C,CH2), 26.85 (2C’s,2xCH2),51.84
(2C’s,2xCH2),94.83(CH=),120.80(Ar-C),127.81(Ar-C),129.32(Ar-C),130.09(Ar-C),132.11(Ar-
C),136.79(Ar-C),140.35(Ar-C),143.82(Ar-C),144.99(=C-N),165.01(C=O), 169.34(C=O), MS
m/z : 531(M+), 532(M+1).
N'-{(E)-[4-(piperidin-1-yl)-1-(4-fluorophenyl)-2, 5-dioxo-2,5-dihydro-1H-pyrrol-3-
yl]methylidene}-4-methylbenzene-1-sulfonohydrazide (5d)
1H NMR(500 MHz. DMSOd6) (δ-ppm) 1.53(br,4H, J=4Hz,2xCH2), 1.59(br,2H, J=4.5Hz,CH2),
2.36(s,3H, CH3), 3.69(br,4H, 2xCH2), 7.27-7.35(m,4H, Ar-H), 7.42(d,2H,J=8Hz, Ar-
H),7.70(t,2H,J=7Hz,Ar-H), 7.81(s,1H, CH=N), 11.27(s,1H,NH), 13C NMR (500MHz, DMSOd6)
20.86 (CH3), 22.95 (1C,CH2), 26.26 (2C’s, 2xCH2), 51.20 (2C’s,2xCH2), 94.13(CH=),115.44
(J=90Hz,Ar-C), 127.21(Ar-C), 127.88(Ar-C), 129 (J=35Hz,Ar-C),129.49 (Ar-C),136.19 (Ar-
C),139.81 (Ar-C),143.21(Ar-C), 144.34 (=C-N),164.68 (C=O), 169.04 (C=O), MS m/z :
470(M+), 471(M+1).
4-methyl-N'-{(E)-[4-(piperidin-1-yl)-1-(4-methylphenyl)-2, 5-dioxo-2, 5-dihydro-1H-pyrrol-3-
yl]methylidene}benzene-1-sulfonohydrazide (5e)
1H NMR(500 MHz. DMSOd6) (δ-ppm) 1.53(br,4H, J=4Hz,2xCH2), 1.58(br,2H,J=5Hz,CH2),
2.32(s,3H, CH3),2.36(s,3H,CH3) 3.69(bs,4H, 2xCH2), 7.15(q,2H,Ja=2Hz and Jb=6.5Hz,Ar-H),
7.24(d,2H,J=8.5Hz,Ar-H), 7.42(d,2H,J=7Hz,Ar-H), 7.71(d,2H,J=8Hz,Ar-H),7.81(s,1H, CH=N),
11.25(s,1H,NH),13CNMR(500MHz,DMSOd6)20.70(J=155Hz,CH3),22.97(1C,CH2),26.25(2C’s,2
xCH2),51.17(2C’s,2xCH2),94.07(CH=),126.71(Ar-C),127.21(Ar-C),129.01(Ar-C),129.49(Ar-
C),136.20(Ar-C),1363.93(Ar-C),139.88(Ar-C),143.20(Ar-C),144.31(=C-N),164.79(C=O),
169.21(C=O), MS m/z : 466(M+), 467(M+1).
N'-{(E)-[4-(piperidin-1-yl)-1-(4-nitrophenyl)-2,5-dioxo-2,5-dihydro-1H-pyrrol-3-
yl]methylidene}-4-methylbenzene-1-sulfonohydrazide (5f)
1H NMR(500 MHz. DMSOd6)
(δ-ppm) 1.60(br,4H, J=5Hz,2xCH2), 2.38(s,3H, CH3),
3.32(d,2H,J=12Hz, CH2), 3.69 (br,4H,2xCH2), 7.43 (d,2H,J=8Hz, Ar-H), 7.63
(q,2H,Ja=2HzandJb=7Hz, Ar-H),7.71(d,2H,J=8Hz,Ar-H), 7.83(s,1H, CH=N),8.33(q,2H,Ja=2Hz

and Jb= 7Hz,Ar-H), 11.33(s,1H,NH), 13C NMR (500MHz, DMSOd6) 20.88 (CH3),22.94
(1C,CH2),26.28 (2C’s,2xCH2),51.37 (2C’s,2xCH2),94.59 (CH=),123.92 (Ar-C),126.74 (Ar-
C),127.24 (Ar-C),127.92 (Ar-C),129.54 (Ar-C),136.18 (Ar-C),137.55 (Ar-C),139.57 (Ar-
C),143.28 (Ar-C),144.53 (Ar-C),145.43 (=C-N),164.12 (C=O), 168.30 (C=O), MS m/z :
497(M+), 498(M+1).
N'-{(E)-[4-(dimethylamino)-2,5-dioxo-1-phenyl-2,5-dihydro-1H-pyrrol-3-yl]methylidene}-4-
methylbenzene-1-sulfonohydrazide (5g)
1H NMR(500 MHz. DMSOd6)
(δ-ppm) 2.38(s,3H,-CH3), 3.23(s,6H,-N(CH3)2),
7.28(q,2H,Ja=1Hz and Jb=7.5Hz,Ar-H), 7.41-7.45(m,5H,Ar-H), 7.72(d,2H,J=8Hz,Ar-H),
7.84(s,1H,-CH=N), 11.27(s,1H,-NH-),13C NMR (500MHz, DMSOd6) 21.48(CH3),43.92(2C’s,
N(CH3)2),94.31(CH=),120.81(Ar-C),127.45(Ar-C),127.76(Ar-C),128.04
(Ar-C),
129.20
(d.J=75Hz,Ar-C), 130.12 (Ar-C), 131.55 (Ar-C),132.18 (d,J=45Hz, Ar-C), 136.77 (Ar-C),140.35
(d,J=60Hz,Ar-C),143.85 (Ar-C), 145.98 (=C-N), 165.08 (C=O), 169.61 C=O), MS m/z : 412
(M+),413 (M+1),414 (M+2).
N'-{(E)-[1-(4-chlorophenyl)-4-(dimethylamino)-2,5-dioxo-2,5-dihydro-1H-pyrrol-3-
yl]methylidene}-4-methylbenzene-1-sulfonohydrazide (5h)
1H NMR(500 MHz. DMSOd6)
(δ-ppm) 2.38(s,3H,-CH3), 3.22(s,6H,-N(CH3)2),
7.32(q,2H,Ja=2Hz and Jb=6.5Hz,Ar-H), 7.40(d,2H,J=8Hz,Ar-H), 7.52(q,2H,Ja=2.5Hz and
Jb=7Hz,Ar-H),7.71(q,2H,Ja=1.5Hz and Jb=6.5Hz,Ar-H) 7.82(s,1H,-CH=N),11.27(s,1H,-NH-
),13CNMR(500MHz,DMSOd6)20.87(CH3),43.31(2C’s,N(CH3)2),93.12(CH=),127.14
(Ar-
C),128.42 (Ar-C),128.58 (Ar-C),129.51 (Ar-C),130.50 (Ar-C),131.77 (Ar-C),136.13 (Ar-
C),139.66 (Ar-C),143.25 (Ar-C),145.36 (=C-N), 164.22 (C=O), 168.70 (C=O), MS m/z : 446
(M+),447 (M+1).
N'-{(E)-[1-(4-bromophenyl)-4-(dimethylamino)-2,5-dioxo-2,5-dihydro-1H-pyrrol-3-
yl]methylidene}-4-methylbenzene-1-sulfonohydrazide (5i)
1H NMR(500 MHz. DMSOd6) (δ-ppm) 2.38(s,3H,-CH3), 3.21(s,6H,-N(CH3)2),
7.26(q,2H,Ja=2Hz and Jb=6.5Hz,Ar-H), 7.41(d,2H,J=8Hz,Ar-H), 7.65(q,2H,Ja=2Hz and
Jb=6.5Hz,Ar-H),7.71(q,2H,Ja=1.5Hz and Jb=6.5Hz,Ar-H) 7.82(s,1H,-CH=N), 11.28(s,1H,-NH-

),13C
NMR(500MHz,DMSOd6)20.89(CH3),43.50(2C’s,
N(CH3)2),93.75(CH=),120.22(Ar-
C),127.17(Ar-C),128.72(Ar-C),129.53(Ar-C),130.95(Ar-C),131.54(Ar-C),136.15(Ar-
C),139.68(Ar-C),143.27(Ar-C),145.39(=C-N),164.19(C=O), 168.66(C=O), MS m/z
490(M+),491(M+1).
:
N'-{(E)-[4-(dimethylamino)-1-(4-fluorophenyl)-2,5-dioxo-2,5-dihydro-1H-pyrrol-3-
yl]methylidene}-4-methylbenzene-1-sulfonohydrazide (5j)
1H NMR(500 MHz. DMSOd6)
(δ-ppm) 2.38(s,3H,-CH3), 3.21(s,6H,-N(CH3)2),
7.26(q,2H,Ja=2Hz and Jb=6.5Hz,Ar-H), 7.41(d,2H,J=8Hz,Ar-H), 7.65(q,2H,Ja=2Hz and
Jb=6.5Hz,Ar-H),7.70(t,2H,J=7Hz ,Ar-H) 7.82(s,1H,-CH=N), 11.27(s,1H,-NH-), 13C NMR
(500MHz,DMSOd6)20.87(CH3),43.20(2C’s,N(CH3)2),93.73(CH=),120.20(Ar-C),127.15(Ar-
C),128.70(Ar-C),129.51(Ar-C),130.94(Ar-C),131.52(Ar-C),136.13(Ar-C),139.66(Ar-
C),143.25,145.36(=C-N),164.17(C=O),168.65(C=O), MS m/z : 490(M+),491(M+1).
4-methyl-N'-{(E)-[4-(dimethylamino)-1-(4-methylphenyl)-2,5-dioxo-2,5-dihydro-1H-pyrrol-3-
yl]methylidene}benzene-1-sulfonohydrazide (5k)
1H NMR(500 MHz. DMSOd6) (δ-ppm) 2.32(s,3H,-CH3),2.38(s,3H,CH3) 3.21(br,6H,-N(CH3)2),
7.14(q,2H,Ja=2Hz and Jb=6.5Hz,Ar-H), 7.24(d,2H,J=8Hz,Ar-H), 7.41(d,2H,J=8Hz,Ar-
H),7.71(d,2H,J=8Hz
,Ar-H)
7.82(s,1H,-CH=N),
11.25(s,1H,-NH-),13C
NMR
(500MHz,DMSOd6)20.54(CH3),20.87(CH3),43.10(2C’s,
N(CH3)2),93.61(CH=),126.69(Ar-
C),127.15(Ar-C),129.01(d,J=15Hz),129.51(Ar-C),136.15(Ar-C),136.94(Ar-C),139.80(Ar-
C),143.23(Ar-C),145.26(=C-N),164.55(C=O), 169.12 (C=O), MS m/z : 426(M+),427(M+1).
N'-{(E)-[4-(dimethylamino)-1-(3-nitrophenyl)-2,5-dioxo-2,5-dihydro-1H-pyrrol-3-
yl]methylidene}-4-methylbenzene-1-sulfonohydrazide (5l)
1H NMR(500 MHz. DMSOd6)
7.41(d,2H,J=8.05Hz,Ar-H),
(δ-ppm) 2.39(s,3H,-CH3), 3.24(s,6H,-N(CH3)2),
7.72(d,2H,J=8.25Hz,Ar-H),
7.76(t,1H,J=7.95Hz,Ar-H),
7.80(q,1H,Ja=1.4Hz andJb=3Hz,Ar-H), 7.84(s,1H,-CH=N), 8.23(m,2H,Ar-H), 11.33(s,1H,-NH-
),13C NMR (500MHz, DMSOd6) 20.86(CH3),43.12(2C’s,N(CH3)2), 93.85(CH=), 120.21(Ar-
C),126.87(Ar-C),127.21(Ar-C),128.73(d,
J=100Hz,Ar-C),
130.98(Ar-C),
131.51(Ar-C),

136.16(Ar-C), 139.80(d,J=55Hz, Ar-C), 143.22, 144.40(=C-N), 164.41(C=O), 168.76(C=O), MS
m/z : 457(M+),458 (M+1).
4.4 Computational Details
Density functional theory calculation was carried out on Intel ® Core™ i7 computer using
Gaussian® 09W, Rev: A.02 Front: 6. 1 version software. Geometry of the synthesized molecule
5c was optimized by the B3LYP functional and 6-311+G(d,P) basis set. The geometrical
parameter, thermodynamic properties, molecular electrostatic potential [MEP], and Mulliken
charges were estimated using the same basis set. Vibrational frequencies were assigned with the
help of Gauss View 5.0 molecular visualization program
4.5. Biological evaluation
The antimicrobial activity of the synthesized derivative was examined by the well diffusion
method. The antibacterial activity was investigated against gram-negative E. coli (ATCC8739)
and gram-positive S. aureus (ATCC25923) bacteria. The antifungal activity was tested for A.
niger (ATCC10231) and C. albicans (ATCC6275). The 0.1 mL suspension of each bacterial
culture (108CFUs/mL) and 0.1 mL spore suspension (108spore/mL) of each fungus was spread
on the sterile Muller Hinton agar plates and Czapek Dox plates respectively. The plates were
allowed to dry for 30 minutes in the laminar airflow and wells were made aseptically using
sterile 6 mm diameter cork borer. The solution of the synthesized derivative was prepared in
DMSO at a concentration of 20 µg mL-1. The wells were filled with 0.3 mL solution of the
derivative. For comparison, the same volume of DMSO was also loaded as a control on a
separate plate. Bacterial plates were incubated for 24 h at 37 °C in the incubator. The fungal plate was
o
incubated for 48 h at 30 C. The experiments were performed in triplicate manner and zone
inhibition was measured by applying a standard deviation method.

Acknowledgments:
The author thanks MVP Samaj Nashik, Principal, K.T.H.M. College, Nashik, and Principal,
K.K.W. College Pimpalgaon Baswant for providing infrastructure facilities. AMB is thankful to
Dr. Keshao Mahale for his help to initiate this work and encouragement. Authors express their
sincere thanks to Savitribai Phule Pune University, Pune for financial support through a minor
research project. We also thank SAIF, Panjab University Chandigarh, and CIF, S.P.P.U. Pune for
characterization help.
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HIGHLIGHTS
 Maleimides (5a-l) were successfully synthesized by condensation of N-aryl 3- and 4-
substituted maleimides with p-toluene sulphonyl hydrazide in ethanol in the acidic
medium at room temperature.
 The novelty of this work is the synthesis was performed at mild condition i.e. room
temperature than other reports.
 The synthesized compound showed good antibacterial activity against Escherichia coli
and Staphylococcus aureus bacteria.
 They also showed efficient antifungal activity towards Aspergillus niger and Candida
albicans fungi.
 We assume that due to good yield, high purity and efficient antimicrobial activity of
synthesized compounds can be used for different biological applications.
 The values of densities, viscosities and relative viscosities of 6-(4-chlorophenyl)-1, 2, 3,
4-tetrahydro-4-oxo-2-thioxopyrimidine-5-carbonitrile in 60% DMSO in the temperature
range 298 to 313 K are studies
 From relative velocity change in Gibb’s free energy, enthalpy and entropy is evaluated.