Synthesis and antimicrobial activity of 2-methyl-5-nitroaniline derivatives: A structure-activity relationship study

Article abstract of DOI:10.1002/cjoc.201190034

A series of 2-methyl-5-nitroaniline derivatives, 5a-5k and 6a-6f, were synthesized in order to determine their in vitro antimicrobial activity. The chemical structures of the synthesized compounds were confirmed by elemental analyses, UV-visible, FT-IR, ¹H NMR and ¹³C NMR spectral studies. The structure-activity relationships of the synthesized compounds were also discussed. Among the synthesized compounds, 5f, 5d, 6b and 6e showed good antimicrobial activity compared to standard drugs. A series of novel 2-methyl-5-nitroaniline derivatives were synthesized and their antimicrobial efficacy was tested. Among the synthesized compounds, 5f and 5d showed good antimicrobial activity. Copyright

Full text of DOI:10.1002/cjoc.201190034

FULL PAPER
Synthesis and Antimicrobial Activity of 2-Methyl-5-nitroaniline
Derivatives: A Structure-Activity Relationship Study
Lingappa, Malleshaa
Kikkeri, N. Mohana*,a
Sreedharamurthy, Satishb
Devaraju, Rakshithb
a Department of Studies in Chemistry, University of Mysore, Manasagangotri, Mysore 570006, India
b Department of Studies in Microbiology, University of Mysore, Manasagangotri, Mysore 570006, India
A series of 2-methyl-5-nitroaniline derivatives, 5a— 5k and 6a— 6f, were synthesized in order to determine
their in vitro antimicrobial activity. The chemical structures of the synthesized compounds were confirmed by ele-
mental analyses, UV-visible, FT-IR, 1H NMR and 13C NMR spectral studies. The structure-activity relationships of
the synthesized compounds were also discussed. Among the synthesized compounds, 5f, 5d, 6b and 6e showed
good antimicrobial activity compared to standard drugs.
Keywords 2-methyl-5-nitroaniline, aldehydes, ketones, antimicrobials
Introduction
have been studied.13,14 Antimicrobial and antifungal ac-
tivities of various Schiff bases have also been re-
ported.15-17 Many Schiff bases are known to be medici-
nally important and are used to design medicinal com-
pounds.18
Diseases caused by microbial infection are a serious
menace to the health of human beings and often have
connection to some of the other diseases, whenever the
body system gets debilitated. Developing antimicrobial
drugs and maintaining their potency, in opposition to
resistance by different classes of microorganisms as well
as a broad spectrum of antibacterial activity are some of
the major concern of research in this area. The com-
pounds containing an azomethine group (CH＝N) are
important in elucidating the mechanism of transamin-
ation and racemisation reactions in biological systems.1,2
Due to the great flexibility and diverse structural aspects,
a wide range of Schiff bases have been synthesized and
their complexation behaviors have been studied.3 They
have been synthesized from a variety of compounds,
such as amino thiazoles, 2-hydroxy-1-napthalaniline,
amino sugars, aromatic aldehydes, ketones, isatin, tria-
zole ring, thiosemicarbazides, amino acids, pyrazolone,
etc.4,5 Antibacterial, antifungal, antitumor and anticancer
activities of Schiff bases have been reported,6,7 and they
are active against a wide range of organisms. Anti-
bacterial activity has been studied more than antifungal
activity, because bacteria can achieve resistance to anti-
biotics through biochemical and morphological modifi-
cations.8,9 Some Schiff bases bearing aryl groups or het-
erocyclic residues possess excellent biological activities
and have attracted the attention of many researchers in
recent years.10-12 The Schiff bases formed from aromatic
aldehydes or aromatic ketones and their derivatives are
quite stable. Due to the great flexibility and diverse
structural aspects of Schiff bases, a wide range of these
compounds have been synthesized and their activities
2-Methyl-5-nitroaniline is used for the preparation of
imatinib which is an anti-cancer agent, and it is cur-
rently marketed as Gleevec. It has also been found to be
effective in the treatment of gastrointestinal stromal tu-
mors (GISTs).19 This selective inhibition of Bcr-Abl
kinase by imatinib has been a successful therapeutic
strategy for chronic myeloid leukemia because of the
high efficacy and mild side effects of this compound.20
2-Methyl-5-nitroaniline passed the animal data screen,
underwent a preliminary toxicological evaluation, and is
being brought to the carcinogen identification committee
for consultation. This is a compilation of the relevant
studies identified during the preliminary toxicological
evaluation.21 The antimicrobial activity of substituted
aniline have been reported.22,23 In connection with such
studies, the present paper reports for the first time the
synthesis of 2-methyl-5-nitroaniline derivatives, 5a—5k
and 6a—6f which are formed during the reaction of
2-methyl-5-nitroaniline (2) with different aldehydes,
3a—3k and different ketones, 4a—4f (Scheme 1).
These synthesized compounds were characterized by
1
elemental analyses, UV-visible, FT-IR, H NMR and
13C NMR studies. Antibacterial and antifungal activities
were reported and structural activity relationship was
also discussed in this paper.
Experimental
All solvents and reagents were purchased from
*
E-mail: knmsvp@yahoo.com; Tel.: 0091-821-2419654; Fax: 094496-27573
Received April 9, 2010; revised September 9, 2010; accepted September 16, 2010.
102
© 2011 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Chin. J. Chem. 2011, 29, 102— 108

Synthesis and Antimicrobial Activity of 2-Methyl-5-nitroaniline Derivatives
Scheme 1 Synthesis of target molecules 5a— 5k and 6a— 6f
Sigma Aldrich Chemicals Pvt Ltd. Melting range was
determined by Veego Melting Point VMP III apparatus.
Elemental analyses were recorded on VarioMICRO su-
peruser V1.3.2 Elementar. The UV-visible spectra were
recorded on Analytik Jena Specord 50 UV-vis spectro-
photometer. The FT-IR spectra were recorded using
KBr discs on FT-IR Jasco 4100 infrared spectro-
Table 1 Chemical structures of the compounds 5a— 5k and
6a— 6f
Compound
R
Compound
R'
5a
6a
1
photometer. H NMR spectra were recorded on Bruker
5b
6b
DRX-400 MHz spectrometer and TMS as an internal
standard. 13C NMR spectra were recorded on Bruker
DRX-100 MHz. The target key intermediate, 2-methyl-
5-nitroaniline (2) was prepared according to literature.24
The reactions of compound 2 with different aldehydes
and ketones were carried out in the presence of absolute
ethanol as a solvent. Synthesized compounds were
characterized by elemental analyses, UV-visible, FT-IR,
1H NMR and 13C NMR. Compounds were purified by
recrystalization method using ethanol. The chemical
structures of all the synthesized compounds are tabu-
lated in Table 1.
5c
5d
5e
6c
6d
6e
5f
6f
General procedure for the synthesis of 2-methyl-5-
nitroaniline derivatives, 5a— 5k and 6a— 6f
Equimolar concentrations of different aldehydes/
ketones (0.01 mol) and 2-methyl-5-nitroaniline (0.01
mol) were stirred for 6 h at room temperature using ab-
solute ethanol (25 mL). The progress of the reaction was
followed by TLC until the reaction was complete. It was
cooled to 0 ℃, the precipitate was filtered, washed with
diethyl ether and the residue was recrystallized from
methanol.
5g
5h
5i
2-Methyl-N-methylene-5-nitroaniline (5a)
The
product was obtained from 2-methyl-5-nitroaniline (2)
(1.53 g, 0.01 mol) and formaldehyde (3a) (0.31 g, 0.01
mol). Yield 86%, m.p. 173—175 ℃; UV-vis (EtOH)
λmax: 510 nm; 1H NMR (DMSO-d6, 400 MHz) δ: 7.71 (s,
2H, H2C＝N), 7.36 (d, J＝1.2 Hz, 1H, Ar-H), 7.16 (d,
J＝5.3 Hz, 1H, Ar-H), 6.40 (s, 1H, Ar-H), 2.14 (s, 3H,
CH3); 13C NMR (DMSO-d6, 100 MHz) δ: 162.8, 162.1,
147.6, 146.2, 136.5, 131.0, 119.3, 84.6; FT-IR (KBr) ν:
3012 (Ar-H), 1613 (C＝N), 1520 (C＝C), 1508, 1345
5j
5k
N-Ethylidene-2-methyl-5-nitroaniline (5b) The
product was obtained from 2-methyl-5-nitroaniline (2)
(1.53 g, 0.01 mol) and acetaldehyde (3b) (0.45 g, 0.01
mol). Yield 61%, m.p. 184—186 ℃; UV-vis (EtOH)
－
1
(NO2), 1260 (C—N) cm . Anal. calcd for C8H8N2O2: C
58.53, H 4.91, N 17.06; found C 58.19, H 4.80, N 16.91.
Chin. J. Chem. 2011, 29, 102— 108
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www.cjc.wiley-vch.de
103

Lingappa et al.
FULL PAPER
λmax: 515 nm; 1H NMR (DMSO-d6, 400 MHz) δ: 8.54 (q,
J＝10.8 Hz, 1H, HC＝N), 7.97 (d, J＝2.0 Hz, 1H,
Ar-H), 7.93 (d, J＝5.2 Hz, 1H, Ar-H), 7.85 (s, 1H,
Ar-H), 3.82 (d, J＝1.1 Hz, 3H, CH3), 2.37 (s, 3H, CH3);
13C NMR (DMSO-d6, 100 MHz) δ: 163.8, 148.0, 146.5,
136.7, 130.9, 119.4, 116.8, 94.4, 45.8; FT-IR (KBr) ν:
3010 (Ar-H), 1635 (C＝N), 1522 (C＝C), 1462, 1376
4-hydroxy-3-methoxybenzaldehyde (3f) (1.53 g, 0.01
mol). Yield 66%, m.p. 103—105 ℃; UV-vis (EtOH)
λmax: 497 nm; 1H NMR (DMSO-d6, 400 MHz) δ: 9.84 (s,
1H, OH), 8.46 (s, 1H, HC＝N), 7.82 (s, 1H, Ar-H), 7.50
(s, 1H, Ar-H), 7.42 (d, J＝5.2 Hz, 1H, Ar-H), 7.28 (d,
J＝10.4 Hz, 1H, Ar-H), 7.14 (d, J＝6.8 Hz, 1H, Ar-H),
6.90 (d, J＝8.0 Hz, 1H, Ar-H), 3.82 (s, 3H, OCH3), 2.37
(s, 3H, CH3); 13C NMR (DMSO-d6, 100 MHz) δ: 163.6,
152.7, 146.5, 136.4, 133.3, 130.9, 129.6, 128.0, 120.2,
116.6, 97.8, 64.2; FT-IR (KBr) ν: 3407 (O—H), 3012
(Ar-H), 1628 (C＝N), 1509 (C＝C), 1462, 1376 (NO2),
1285 (C—N), 1083 (C—O) cm－ 1. Anal. calcd for
C15H14N2O4: C 62.93, H 4.93, N 9.79; found C 62.71, H
4.73, N, 10.01.
－
1
(NO2), 1303 (C—N) cm . Anal. calcd for C9H10N2O2: C
60.66, H 5.66, N 15.72; found C 60.39, H 5.41, N 15.91.
N-Benzylidene-2-methyl-5-nitroaniline (5c) The
product was obtained from 2-methyl-5-nitroaniline (2)
(1.53 g, 0.01 mol) and benzaldehyde (3c) (1.10 g, 0.01
mol). Yield 67%, m.p. 152—154 ℃; UV-vis (EtOH)
λmax: 529 nm; 1H NMR (DMSO-d6, 400 MHz) δ: 9.00 (s,
1H, HC＝N), 8.16 (s, 1H, Ar-H), 8.04 (d, J＝8.2 Hz, 2H,
Ar-H), 7.73 (d, J＝2.4 Hz, 1H, Ar-H), 7.58 (d, J＝6.0
Hz, 1H, Ar-H), 7.44 (t, J＝15.9 Hz, 2H, Ar-H), 7.00 (t,
J＝7.3 Hz, 1H, Ar-H), 2.47 (s, 3H, CH3); 13C NMR
(DMSO-d6, 100 MHz) δ: 163.1, 153.1, 146.7, 136.7,
133.9, 131.4, 129.3, 127.8, 119.8, 116.9, 94.5; FT-IR
(KBr) ν: 3004 (Ar-H), 1635 (C＝N), 1525 (C＝C), 1463,
1376 (NO2), 1303 (C — N) cm － 1. Anal. calcd for
C14H12N2O2: C 69.99, H 5.03, N 11.66; found C 70.12,
H 4.91, N 11.71.
N-(2-Chloro-6-fluorobenzylidene)-2-methyl-5-
nitroaniline (5g) The product was obtained from
2-methyl-5-nitroaniline (2) (1.53 g, 0.01 mol) and
2-chloro-6-fluorobenzaldehyde (3g) (1.59 g, 0.01 mol).
Yield 64%, m.p. 113—115 ℃; UV-vis (EtOH) λmax
:
495 nm; 1H NMR (DMSO-d6, 400 MHz) δ: 8.53 (s, 1H,
HC＝N), 7.97 (t, J＝8.7 Hz, 1H, Ar-H), 7.91 (d, J＝6.1
Hz, 1H, Ar-H), 7.84 (s, 1H, Ar-H), 7.50 (d, J＝2.0 Hz,
1H, Ar-H), 7.06 (s, 1H, Ar-H), 7.04 (d, J＝5.2 Hz, 1H,
Ar-H), 2.36 (s, 3H, CH3); 13C NMR (DMSO-d6, 100
MHz) δ: 164.7, 161.3, 152.3, 136.4, 130.3, 129.4, 124.5,
120.1, 118.5, 114.3, 97.8, 64.2; FT-IR (KBr) ν: 3037
(Ar-H), 1638 (C＝N), 1516 (C＝C), 1455, 1349 (NO2),
N-(4-Methoxybenzylidene)-2-methyl-5-nitroaniline
(5d) The product was obtained from 2-methyl-5-
nitroaniline (2) (1.53 g, 0.01 mol) and 4-methoxyben-
zaldehyde (3d) (1.23 g, 0.01 mol). Yield 64%, m.p.
－
1
1310 (C—N), 1253 (C—F), 738 (C—Cl) cm . Anal.
calcd for C14H10ClFN2O2: C 57.45, H 3.44, N 9.57;
found C 57.31, H 3.73 N 9.37.
1
173—175 ℃; UV-vis (EtOH) λmax: 520 nm; H NMR
(DMSO-d6, 400 MHz) δ: 8.46 (s, 1H, HC＝N), 7.56 (s,
1H, Ar-H), 7.28 (d, J＝1.8 Hz, 1H, Ar-H), 7.20 (d, J＝
3.5 Hz, 2H, Ar-H), 7.14 (d, J＝5.1 Hz, 1H, Ar-H), 6.91
(d, J＝4.7 Hz, 2H, Ar-H), 3.41 (s, 3H, OCH3), 2.37 (s,
3H, CH3); 13C NMR (DMSO-d6, 100 MHz) δ: 163.2,
152.9, 146.7, 136.5, 134.0, 131.3, 129.2, 127.8, 119.7,
116.8, 94.5; FT-IR (KBr) ν: 3094 (Ar-H), 1634 (C＝N),
1522 (C＝C), 1464, 1352 (NO2), 1306 (C—N), 1130
N-(2-Chlorobenzylidene)-2-methyl-5-nitroaniline
(5h) The product was obtained from 2-methyl-5-
nitroaniline (2) (1.53 g, 0.01 mol) and 2-chlorobenzal-
dehyde (3h) (1.41 g, 0.01 mol). Yield 61%, m.p. 122—
124 ℃ ; UV-vis (EtOH) λmax: 490 nm; 1H NMR
(DMSO-d6, 400 MHz) δ: 8.55 (s, 1H, HC＝N), 7.97 (t,
J＝11.3 Hz, 1H, Ar-H), 7.91 (d, J＝3.4 Hz, 1H, Ar-H),
7.89 (d, J＝4.1 Hz, 1H, Ar-H), 7.83 (s, 1H, Ar-H), 7.41
(d, J＝2.7 Hz, 1H, Ar-H), 7.35 (d, J＝5.0 Hz, 1H, Ar-H),
7.31 (d, J＝6.2 Hz, 1H, Ar-H), 2.37 (s, 3H, CH3);
13C NMR (DMSO-d6, 100 MHz) δ: 164.2, 161.5, 154.2,
135.4, 130.3, 129.3, 124.2, 120.0, 118.3, 114.1, 97.6,
64.1; FT-IR (KBr) ν: 3045 (Ar-H), 1618 (C＝N), 1509
(C＝C), 1463, 1376 (NO2), 1348 (C—N), 739 (C—Cl)
－
1
(C—O) cm . Anal. calcd for C15H14N2O3: C 66.66, H
5.22, N 10.36; found C 66.49, H 5.27, N 10.41.
2-[(2-Methyl-5-nitrophenylimino)methyl]phenol
(5e) The product was obtained from 2-methyl-5-
nitroaniline (2) (1.53 g, 0.01 mol) and 2-hydroxyben-
zaldehyde (3e) (1.23 g, 0.01 mol). Yield 71%, m.p. 100—
103 ℃ ; UV-vis (EtOH) λmax: 527 nm; 1H NMR
(DMSO-d6, 400 MHz) δ: 9.34 (s, 1H, O—H), 8.72 (s,
1H, HC＝N), 8.04 (d, J＝6.2 Hz, 1H, Ar-H), 7.86 (s, 1H,
Ar-H), 7.61—7.53 (m, 2H, Ar-H), 7.48 (d, J＝3.5 Hz, 1H,
Ar-H), 7.38 (d, J＝2.7 Hz, 1H, Ar-H), 7.36 (d, J＝4.8 Hz,
1H, Ar-H), 2.36 (s, 3H, CH3); 13C NMR (DMSO-d6, 100
MHz) δ: 163.2, 152.9, 146.7, 136.6, 133.8, 131.4, 129.3,
127.8, 119.8, 116.8, 94.6; FT-IR (KBr) ν: 3367 (O—H),
3041 (Ar-H), 1618 (C＝N), 1509 (C＝C), 1463, 1376
－
1
cm . Anal. calcd for C14H11ClN2O2: C 61.21, H 4.04, N
10.20; found C 60.97, H 4.11, N 10.13.
N-(4-Fluorobenzylidene)-2-methyl-5-nitroaniline
(5i) The product was obtained from 2-methyl-5-
nitroaniline (2) (1.53 g, 0.01 mol) and 4-fluorobenzal-
dehyde (3i) (1.25 g, 0.01 mol). Yield 74%, m.p. 110—
112 ℃ ; UV-vis (EtOH) λmax: 494 nm; 1H NMR
(DMSO-d6, 400 MHz) δ: 8.60 (s, 1H, HC＝N), 7.95 (d,
J＝10.3 Hz, 2H, Ar-H), 7.90 (d, J＝5.6 Hz, 2H, Ar-H),
7.83 (s, 1H, Ar-H), 7.51 (d, J＝1.7 Hz, 1H, Ar-H), 7.06
(d, J＝5.2 Hz, 1H, Ar-H), 2.36 (s, 3H, CH3); 13C NMR
(DMSO-d6, 100 MHz) δ: 164.1, 161.3, 153.9, 135.4,
130.3, 129.2, 124.1, 120.0, 118.4, 114.5, 97.6, 64.3;
FT-IR (KBr) ν: 3017 (Ar-H), 1631 (C＝N), 1510 (C＝
－
1
(NO2), 1285 (C—N) cm . Anal. calcd for C14H12N2O3:
C 65.62, H 4.72, N 10.93; found C 65.71, H 4.73, N,
10.77.
2-Methoxy-4-[(2-methyl-5-nitrophenylimino)meth-
yl]phenol (5f)
The product was obtained from
2-methyl-5-nitroaniline (2) (1.53 g, 0.01 mol) and
104
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Synthesis and Antimicrobial Activity of 2-Methyl-5-nitroaniline Derivatives
－
1
C), 1462, 1371 (NO2), 1345 (C—N), 1285 (C—F) cm .
Anal. calcd for C14H11FN2O2: C 65.11, H 4.29, N 10.85;
found C 65.31, H 4.23, N 10.77.
Hz, 1H, Ar-H), 7.32 (d, J＝5.6 Hz, 1H, Ar-H), 3.82 (s,
3H, CH3), 2.36 (s, 3H, CH3); 13C NMR (DMSO-d6, 100
MHz) δ: 164.2, 161.3, 153.8, 134.6, 131.5, 128.2, 125.1,
124.8, 122.2, 120.1, 118.5, 116.2, 114.3, 98.3, 65.5;
FT-IR (KBr) ν: 3367 (N—H), 3010 (Ar-H), 1618 (C＝
N-(4-Ethoxybenzylidene)-2-methyl-5-nitroaniline
(5j) The product was obtained from 2-methyl-5-
nitroaniline (2) (1.53 g, 0.01 mol) and 4-ethoxybenzal-
dehyde (3j) (1.51 g, 0.01 mol). Yield 63%, m.p. 113—
115 ℃ ; UV-vis (EtOH) λmax: 505 nm; 1H NMR
(DMSO-d6, 400 MHz) δ: 8.51 (s, 1H, HC＝N), 7.96 (d,
J＝14.0 Hz, 2H, Ar-H), 7.86 (d, J＝3.7 Hz, 2H, Ar-H),
7.58 (s, 1H, Ar-H), 7.50 (d, J＝1.8 Hz, 1H, Ar-H), 7.54
(d, J＝6.0 Hz, 1H, Ar-H), 4.12 (q, J＝12.3 Hz, 2H,
CH2), 2.36 (s, 3H, CH3), 1.35 (t, J＝8.5 Hz, 3H, CH3);
13C NMR (DMSO-d6, 100 MHz) δ: 164.3, 161.2, 154.0,
134.1, 131.0, 130.2, 128.5, 125.2, 124.3, 120.1, 118.5,
114.2, 98.1, 65.3; FT-IR (KBr) ν: 3024 (Ar-H), 1634
(C＝N), 1578 (C＝C), 1464, 1352 (NO2), 1306 (C—N),
1130 (C—O) cm－ 1. Anal. calcd for C16H16N2O3: C
67.59, H 5.67, N 9.85; found C 67.71, H 5.73, N 10.07.
2-Methyl-5-nitro-N-(pyridin-2-ylmethylene)aniline
(5k) The product was obtained from 2-methyl-5-
nitroaniline (2) (1.53 g, 0.01 mol) and 2-pyridine carbox-
aldehyde (3k) (1.10 g, 0.01 mol). Yield 67%, m.p. 108—
110 ℃ ; UV-vis (EtOH) λmax: 512 nm; 1H NMR
(DMSO-d6, 400 MHz) δ: 8.53 (s, 1H, HC＝N), 7.86 (t,
J＝10.3 Hz, 1H, Ar-H), 7.85 (d, J＝7.1 Hz, 1H, Ar-H),
7.83 (d, J＝5.1 Hz, 1H, Ar-H), 7.62 (s, 1H, Ar-H), 7.60 (t,
J＝9.3 Hz, 1H, Ar-H), 7.57 (d, J＝2.2 Hz, 1H, Ar-H),
7.52 (d, J＝5.1 Hz, 1H, Ar-H), 2.37 (s, 3H, CH3);
13C NMR (DMSO-d6, 100 MHz) δ: 164.1, 161.3, 153.9,
134.1, 131.3, 128.7, 125.2, 122.0, 114.2, 98.1, 65.3;
FT-IR (KBr) ν: 3032 (Ar-H), 1635 (C＝N), 1524 (C＝C),
－
1
N), 1574 (C＝C), 1463, 1348 (NO2), 1285 (C—N) cm .
Anal. calcd for C17H15N3O2: C 69.61, H 5.15, N 14.33;
found C 69.72, H 5.23, N 14.11.
2-Methyl-5-nitro-N-[1-(pyridin-3-yl)ethylidene]-
aniline (6c) The product was obtained from 2-methyl-
5-nitroaniline (2) (1.53 g, 0.01 mol) and 1-(pyridin-3-
yl)ethanone (4c) (1.22 g, 0.01 mol). Yield 64%, m.p.
1
126—128 ℃; UV-vis (EtOH) λmax: 525 nm; H NMR
(DMSO-d6, 400 MHz) δ: 8.53 (s, 1H, Ar-H), 7.97—7.95
(m, 3H, Ar-H), 7.89 (s, 1H, Ar-H), 7.06—7.04 (d, J＝
8.8 Hz, 2H, Ar-H), 3.30 (s, 3H, CH3), 2.36 (s, 3H, CH3);
13C NMR (DMSO-d6, 100 MHz) δ: 164.2, 161.4, 153.5,
134.5, 131.1, 128.3, 125.4, 122.4, 118.7, 114.1, 98.7,
65.4; FT-IR (KBr) ν: 3031 (Ar-H), 1631 (C＝N), 1527
－
1
(C＝C), 1462, 1375 (NO2), 1342 (C—N) cm . Anal.
calcd for C14H13N3O2: C 65.87, H 5.13, N 16.46; found
C 65.72, H 5.23, N 16.21.
2-Methyl-N-[1-(naphthalen-2-yl)ethylidene]-5-
nitroaniline (6d) The product was obtained from
2-methyl-5-nitroaniline (2) (1.53 g, 0.01 mol) and
1-(naphthalen-2-yl)ethanone (4d) (1.71 g, 0.01 mol).
Yield 68%, m.p. 134—136 ℃; UV-vis (EtOH) λmax
:
515 nm; 1H NMR (DMSO-d6, 400 MHz) δ: 8.46 (d, J＝
6.3 Hz, 2H, Ar-H), 7.94 (t, J＝12.0 Hz, 2H, Ar-H), 7.49
(d, J＝8.0 Hz, 2H, Ar-H), 7.42 (s, 1H, Ar-H), 7.38 (d,
J＝7.3 Hz, 1H, Ar-H), 7.12 (d, J＝1.8 Hz, 1H, Ar-H),
6.90 (d, J＝5.2 Hz, 1H, Ar-H), 3.82 (s, 3H, CH3), 2.37
(s, 3H, CH3); 13C NMR (DMSO-d6, 100 MHz) δ: 165.8,
164.5, 161.7, 157.2 153.1, 147.3, 141.4, 134.2, 131.6,
129.3, 124.4, 121.4, 119.7, 116.2, 114.5, 97.4, 66.4;
FT-IR (KBr) ν: 3010 (Ar-H), 1633 (C＝N), 1528 (C＝
－
1
1461, 1373 (NO2), 1341 (C—N) cm . Anal. calcd for
C13H11N3O2: C 64.72, H 4.60, N 17.42; found C 64.91, H
4.73, N 17.17.
2-Methyl-5-nitro-N-(1-phenylethylidene)aniline
(6a) The product was obtained from 2-methyl-5-
nitroaniline (2) (1.53 g, 0.01 mol) and acetophenone (4a)
(1.21 g, 0.01 mol). Yield 76%, m.p. 125—127 ℃;
－
1
C), 1461, 1367 (NO2), 1341 (C—N) cm . Anal. calcd
for C19H16N2O2: C 74.98, H 5.30, N 9.20; found C 75.02,
H 5.21, N 9.01.
1
UV-vis (EtOH) λmax: 510 nm; H NMR (DMSO-d6, 400
2-Methyl-5-nitro-N-[1-(pyridin-4-yl)ethylidene]-
aniline (6e) The product was obtained from 2-methyl-
5-nitroaniline (2) (1.53 g, 0.01 mol) and 1-(pyridin-4-
yl)ethanone (4e) (1.22 g, 0.01 mol). Yield 73%, m.p.
MHz) δ: 7.72 (s, 1H, Ar-H), 7.42 (d, J＝10.0 Hz, 2H,
Ar-H), 7.16 (t, J＝16.1 Hz, 2H, Ar-H), 7.61 (t, J＝10.3
Hz, 1H, Ar-H), 7.55 (d, J＝2.7 Hz, 1H, Ar-H), 7.51 (d,
J＝4.9 Hz, 1H, Ar-H), 3.15 (s, 3H, CH3), 2.14 (s, 3H,
CH3); 13C NMR (DMSO-d6, 100 MHz) δ: 164.3, 161.1,
153.9, 134.1, 131.3, 128.7, 125.2, 124.5, 122.0, 120.5,
114.3, 98.5, 65.7; FT-IR (KBr) ν: 3030 (Ar-H), 1631
(C＝N), 1521 (C＝C), 1467, 1371 (NO2), 1343 (C—N)
1
127—139 ℃; UV-vis (EtOH) λmax: 525 nm; H NMR
(DMSO-d6, 400 MHz) δ: 8.71 (s, 1H, Ar-H), 8.02 (t, J＝
12.1 Hz, 2H, Ar-H), 7.86 (d, J＝1.7 Hz, 1H, Ar-H), 7.60
(t, J＝8.1 Hz, 2H, Ar-H), 7.53 (d, J＝6.1 Hz, 1H, Ar-H),
3.30 (s, 3H, CH3), 2.35 (s, 3H, CH3); 13C NMR
(DMSO-d6, 100 MHz) δ: 164.2, 161.3, 153.2, 134.0,
131.1, 128.3, 125.5, 122.2, 118.4, 114.2, 98.6, 66.4;
FT-IR (KBr) ν: 3032 (Ar-H), 1631 (C＝N), 1526 (C＝
－
1
cm . Anal. calcd for C15H14N2O2: C 70.85, H 5.55, N
11.02; found C 70.71, H 5.73, N 11.17.
N-[1-(1H-indol-3-yl)ethylidene]-2-methyl-5-nitro-
aniline (6b) The product was obtained from 2-methyl-
5-nitroaniline (2) (1.53 g, 0.01 mol) and 1-(1H-indol-3-
yl)ethanone (4b) (1.60 g, 0.01 mol). Yield 72%, m.p.
－
1
C), 1462, 1375 (NO2), 1341 (C—N) cm . Anal. calcd
for C14H13N3O2: C 65.87, H 5.13, N 16.46; found C
65.92, H 5.21, N 16.31.
1
145—148 ℃; UV-vis (EtOH) λmax: 530 nm; H NMR
3-[1-(2-Methyl-5-nitrophenylimino)ethyl]benzoni-
trile (6f) The product was obtained from 2-methyl-5-
nitroaniline (2) (1.53 g, 0.01 mol) and 3-acetylbenzoni-
trile (4f) (1.46 g, 0.01 mol). Yield 71%, m.p. 141—143
(DMSO-d6, 400 MHz) δ: 8.54 (s, 1H, NH), 7.96 (t, J＝
4.0 Hz, 2H, Ar-H), 7.85 (s, 1H, Ar-H), 7.50—7.48 (d,
J＝8.0 Hz, 2H, Ar-H), 7.08 (s, 1H, CH), 7.06 (d, J＝2.1
Chin. J. Chem. 2011, 29, 102— 108
© 2011 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.cjc.wiley-vch.de
105

Lingappa et al.
FULL PAPER
1
℃; UV-vis (EtOH) λmax: 510 nm; H NMR (DMSO-d6,
400 MHz) δ: 8.53 (d, J＝4.8 Hz, 1H, Ar-H), 7.96 (t, J＝
9.3 Hz, 1H, Ar-H), 7.91 (d, J＝7.2 Hz, 1H, Ar-H), 7.84
(s, 1H, Ar-H), 7.56 (d, J＝2.5 Hz, 1H, Ar-H), 7.52 (s,
1H, Ar-H), 7.06 (d, J＝5.1 Hz, 1H, Ar-H), 3.31 (s, 3H,
CH3), 2.36 (s, 3H, CH3); 13C NMR (DMSO-d6, 100
MHz) δ: 165.0, 164.7, 163.4, 154.5, 143.1, 131.5, 128.1,
125.7, 124.8, 122.4, 120.1, 114.2, 96.1, 65.2; FT-IR
(KBr) ν: 3033 (Ar-H), 2745 (C≡N), 1632 (C＝N),
absorption spectra of synthesized compounds show new
bands and appearance of longer wavelength absorption
band in the visible region in UV-visible spectrum con-
firmed the formation of synthesized compounds. The
absorptions around 3000 cm－ in synthesized com-
1
pounds confirm the aromatic stretching vibrations, and
the appearance of a medium to strong absorption bands
above 1600 cm－ due to a stretching vibration of the
1
azomethine (C ＝ N) bond formation in synthesized
compounds via condensation. The proton spectral data
agree with respect to the number of protons and their
chemical shifts with the proposed structures. The proton
spectral data of the key intermediate, 2-methyl-5-
nitroaniline (2) show resonance at δ 5.52 (s, 2H, NH2).
For compound 5a—5k the above resonance disappeared
and additional resonances assigned to the CH＝N (δ
7.71—9.00) were observed, which confirmed the con-
densation between the amino group and carbonyl group.
The 13C NMR spectra presented the correct number of
carbon atoms at the appropriate chemical shift values.
The synthesized compounds showed absorptions at δ
14.3—19.5 due to CH3, δ 110.2—154.3 due to aromatic
carbon atoms, δ 259.7 due to C—O, δ 160.1—163.5 due
to CH＝N groups.
－
1
1527 (C＝C), 1465, 1373 (NO2), 1341 (C—N) cm .
Anal. calcd for C16H13N3O2: C 68.81, H 4.69, N15.05;
found C 69.01, H 4.91, N 15.31.
In vitro antibacterial activity
Antibacterial activity of the synthesized compounds
was determined against gram-positive bacteria (Bacillus
subtilis, Staphylococcus aureus) and gram-negative
bacteria (Xanthomonas campestris and Escherichia coli)
in DMF by disc diffusion method on nutrient agar me-
dium.25 The sterile medium (Nutrient Agar medium, 15
mL) in each petriplates was uniformly smeared with
cultures of Gram ＋ve and Gram －ve bacteria. Sterile
discs of 10 mm diameter (Hi-Media) were made in each
of the petriplates, to which 50 µL (1 mg/mL i.e., 50
µg/disc) of the different synthesized compounds were
added. The treatments also included 50 µL of DMF and
streptomycin as negative and positive control for com-
parison. For each treatment, three replicates were main-
tained. The plates were incubated at (25±2) ℃ for 24
h and the size of the resulting zone of inhibition, if any,
was determined. The data were subjected to analysis of
variance (ANOVA).
Antibacterial activity
The antibacterial activity of compounds, 5a—5k and
6a—6f, were evaluated and compared with streptomycin
as standard drug. Compounds, 5d, 5f, 6b and 6e,
showed good antibacterial properties against four
pathogenic bacterial strains compared with other com-
pounds. Compound 5f exhibit inhibition in the range of
12.8—20.2 mm. Compounds 5d, 6b and 6e exhibit in-
hibition in the range of 12.6—18.5 mm, 12.3—18.4 mm
and 12.1—18.1 mm against B. subtilis, S. aureus, E. coli
and X. campestris, respectively. Compounds 6c, 5k, 5g,
5j and 5e, showed moderate antibacterial activity
against four pathogenic bacterial strains. The other
compounds showed minimal antibacterial activity.
In vitro antifungal activity
The synthesized compounds were screened for their
antifungal activity against Fusarium oxysporum in DMF
by poisoned food technique.26 Potato Dextrose Agar
(PDA) media was prepared and about 15 mL of PDA
was poured into each petriplate and allowed to solidify.
5 mm disc of 7 d old culture of the test fungi was placed
at the center of the petriplates and incubated at 26 ℃
for 7 d. After incubation the percentage inhibition was
measured and three replicates were maintained for each
treatment. Activity of compounds was compared with
standard drug nystatin. All the synthesized compounds
were tested (at the dosage of 500 µL of the novel com-
pounds/petriplate, where concentration was 0.1 mg/mL)
by poisoned food technique.
Antifungal activity
All the synthesized compounds 5a—5k and 6a—6f
were also tested against F. oxysporum and found that
compounds showed varying degree of percentage inhi-
bition. The compounds, 5d, 5f, 6b and 6e, showed good
antifungal activity than other compounds in the series
5a—5k and 6a—6f against tested strain. From the re-
sults, it is evident that most of the compounds are mod-
erately active and few are weakly active. The antimi-
crobial activity results of synthesized compounds were
compared with standard drugs as depicted in Table 2.
Among the compounds, 5a—5k and 6a—6f, showed
antimicrobial activity in the order of 5f＞5d＞6b＞6e＞
6c＞5k＞5g＞5j＞5e＞6d＞6a＞5c＞5b＞5a＞5h＞
6f＞5i against tested bacterial and fungal strains.
Results and discussion
Schiff bases of 2-methyl-5-nitroaniline derivatives,
5a—5k and 6a—6f were synthesized from condensation
reaction of 2-methyl-5-nitroaniline with different alde-
hydes/ketones (Scheme 1). The elemental analyses data
showed good agreement between the experimentally
determined values and the theoretically calculated val-
ues within the limits of permissible error. The electronic
Structure-activity relationships
Initial structure activity relationship can be drawn for
106
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© 2011 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Chin. J. Chem. 2011, 29, 102— 108

Synthesis and Antimicrobial Activity of 2-Methyl-5-nitroaniline Derivatives
the compounds, 5a—5k and 6a—6f. Antimicrobial and
structure activity relationship studies of Schiff bases of
3-amino-2-phenylquinazolin-4(3H)-one have been re-
ported.27 In the present study, different electron with-
drawing and electron donating groups attached to
phenyl ring as substituent were linked to azomethine
group. The close survey of antimicrobial efficacy indi-
cated that the inhibition values of all the compounds
exhibited a varied range (8.0—20.2 mm and 23.4%—
73.1%) of antibacterial and antifungal activities against
all the tested microbial strains. The electron donating
methoxy group of phenyl ring in 5f produces enhanced
activity probably by m-effect compared to the p-position
in 5d. This indicates the positional requirement of
methoxy group on phenyl ring for enhanced activity.
The electron withdrawing chlorine and fluorine atoms in
5g, pyridine ring in 5k, electron donating hydroxyl
group in 5e showed moderate activity whereas the chlo-
rine atom alone in 5h produces weak antimicrobial
activity. The compounds having unsubstituted phenyl
ring in 5c and 6a revealed relatively moderate antim-
icrobial activity. However, 6a showed better activity
compared to 5c probably due to the presence of methyl
group in 6a. The compounds 5a and 5b are weakly
active which may be due to the absence of phenyl ring.
The electron donating ethoxy group in 5j showed mod-
erate antimicrobial activity against tested microbial
strains. The electron withdrawing fluorine atom in 5i
showed weak antimicrobial activity. The indole group in
6b, pyridine group in 6c and 6e produce good anti-
microbial activity. Naphthalene ring in 6d showed mod-
erate activity whereas the benzonitrile group in 6f pro-
duces weak antimicrobial activity. The above SAR cor-
relation studies reveal that, the nature of the linkage
(substituent on aromatic ring) influences the antimicro-
bial activity.
Conclusion
In conclusion, a series of novel 2-methyl-5-nitro-
aniline derivatives, 5a—5k and 6a—6f were synthe-
sized in good yield, characterized by different spectral
studies, and their antimicrobial activities were deter-
mined against clinically important pathogens. Com-
pounds, 5d, 5f, 6b, and 6e demonstrated good inhibition
against bacterial and fungal strains tested. The SAR
studies revealed that the substituents on aromatic ring
are responsible for the antimicrobial activity of these
classes of agents. On the basis of their activity, these
derivatives were identified as viable leads for further
studies.
Acknowledgements
One of the authors Mallesha Lingappa is grateful to
University Grants Commission, New Delhi, for financial
support under UGC-RFSMS scheme, and thank Univer-
sity of Mysore for the award of Junior Research Fel-
lowship.
Table 2 In vitro antibacterial and antifungal activities of compounds 5a— 5k and 6a— 6f
Zone of inhibition in diameter/mm
Inhibition/%
F. oxysporum
25.6
Compound
B. subtilis
11.8
12.0
12.1
17.1
14.2
17.3
14.9
11.5
10.1
14.5
16.0
13.4
17.0
16.1
13.6
16.7
11.0
18.0
—
S. aureus
11.7
11.9
12.5
18.5
15.3
20.2
15.9
11.0
10.2
15.7
17.6
15.0
18.4
18.0
14.3
18.1
10.3
20.9
—
X. campestris
8.8
E. coli
8.3
5a
5b
9.0
8.9
30.5
5c
9.1
9.3
32.3
5d
12.6
10.1
12.8
10.4
8.8
13.9
10.3
14.0
10.6
8.3
72.4
5e
49.0
5f
73.1
5g
68.2
5h
25.3
5i
8.0
8.1
23.4
5j
10.2
12.0
9.2
10.5
13.1
10.1
13.8
13.3
9.6
52.4
5k
68.7
6a
38.7
6b
12.3
12.3
9.4
70.7
6c
58.6
6d
6e
38.9
12.1
8.1
13.5
8.1
70.1
6f
23.7
Streptomycin
Nystatin
13.2
—
14.2
—
—
80
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© 2011 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.cjc.wiley-vch.de
107

Lingappa et al.
FULL PAPER
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Chin. J. Chem. 2011, 29, 102— 108