Design, synthesis and in vitro evaluation of tetrahydropyrimidine-isatin hybrids as potential antitubercular and antimalarial agents

Article abstract of DOI:10.1016/j.cclet.2012.05.004

A series of 5-substituted-3-[{5-(6-methyl-2-oxo/thioxo-4-phenyl-1,2,3,4- tetrahydropyrimidin-5-yl)-1,3,4-oxadiazol-2-yl}-imino]-1,3-dihydro-2H-indol-2- one were synthesized, characterized and screened for their anti-tubercular and antimalarial activity.

Full text of DOI:10.1016/j.cclet.2012.05.004

Available online at www.sciencedirect.com
Chinese Chemical Letters 23 (2012) 785–788
www.elsevier.com/locate/cclet
Design, synthesis and in vitro evaluation of
tetrahydropyrimidine–isatin hybrids as potential
antitubercular and antimalarial agents
*
Tarunkumar Nanjibhai Akhaja, Jignesh Priyakant Raval
Department of Pharmaceutical Chemistry, Ashok & Rita Patel Institute of Integrated Study and Research in Biotechnology and
Allied Sciences (ARIBAS), New Vallabh Vidyanagar 388121, India
Received 7 March 2012
Available online 9 June 2012
Abstract
A series of 5-substituted-3-[{5-(6-methyl-2-oxo/thioxo-4-phenyl-1,2,3,4-tetrahydropyrimidin-5-yl)-1,3,4-oxadiazol-2-yl}-im-
ino]-1,3-dihydro-2H-indol-2-one were synthesized, characterized and screened for their anti-tubercular and antimalarial activity.
# 2012 Jignesh Priyakant Raval. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords: Tetrahydropyrimidines–isatin hybrid; Biginelli reaction; In vitro antitubercular; Antimalarial activity
1,3-Dihydro-2H-indol-2-one nucleus is used as a versatile lead molecule for designing potential antitubercular [1],
antiviral [2], anticonvulsant [3] and anti-tumor [4] agents. Similarly, imine bases of 1,3-dihydro-2H-indol-2-ones were
reported for various biological activities [5–7]. Literature survey revealed that introduction of electron-withdrawing
groups at positions 5, 6, and 7 greatly increased activity from that of 1,3-dihydro-2H-indol-2-one, with substitution at
the 5th position being most favorable. This is not surprising, as C-5 substitution has been associated with increased
biological activity [8,9] and, the presence of substituted aromatic/heteoaromatic ring at 3rd position has been reported
to be associated with antimicrobial properties [10,11]. Also 3-substituted indolin-2-ones have been identified as a
versatile scaffold for the development of protein kinase inhibitors which exhibit selectivity toward different receptor
tyrosine kinases (RTKs) by altering the substituents [12]. Sun et al. reported that several 3-substituted indolin-2-one
derivatives containing bulky groups in the phenyl ring at the C-3 position of indolin-2-ones showed high selectivity
toward the EGF and Her-2 RTKs [13]. The various substituent at 3rd position of the indolin-2-ones reported, were
various substituted phenyl ring [14], heterocyclic ring [15] and aliphatic system [16]. Recently George et al. had also
reported some of the pyrimidinone-oxadiazolylindolinones as promising antimicrobials together with antioxidant
activities [17]. This observations led us to synthesis 5-substituted-3-[{5-(6-methyl-2-oxo/thioxo-4-phenyl-1,2,3,4
tetrahydropyrimidin-5-yl)-1,3,4-oxadiazol-2-yl}imino]-1,3-dihydro-2H-indol-2-ones 4a-l, using 5-(5-amino-1,3,4-
oxadiazol-2-yl)-6-methyl-4-phenyl-3,4-dihydropyrimidin-2(H)-one/thiones and different 5-substituted indoline-2,3-
dione.
* Corresponding author.
E-mail addresses: tarun.pch@gmail.com (T.N. Akhaja), drjpraval@yahoo.co.in (J.P. Raval).
1001-8417/$ – see front matter # 2012 Jignesh Priyakant Raval. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
http://dx.doi.org/10.1016/j.cclet.2012.05.004

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T.N. Akhaja, J.P. Raval / Chinese Chemical Letters 23 (2012) 785–788
C H O C
2
5
2
O
H C
3
CO C H
CONHNH
2
c)
2
2
5
a)
i)
HN
b)
HN
+
NH
2
X
N
H
CH
3
N
H
X
iii)
O
CH
1a/1b
3
H N
2
ii)
X
2a/2b
Here:
X= O / S;
N
N
d)
N
N
R= H, Br, NO , F, I and Cl.
2
a) CaCl 20 mole %,
2
NH
HN
2
O
N
HN
b) NH NH .H O, Conc. H SO ,
R
O
2
2
2
2
4
c) CNBr, EtOH, NaHCO ,
3
X
N
H
3a/3b
CH
3
O
CH
3
d) 5-substituted isatine, MeOH, drops of CH COOH
X
N
H
3
N
H
4a-l
Scheme 1. Synthetic pathway leading to the title compounds 4a-l.
The synthetic pathway leading to the title compounds 4a-l is given in Scheme 1. In a typical experimental
procedure, a solution of b-ketoester, aldehyde and urea/thiourea in ethanol was heated under reflux in the presence of
catalytic amount of CaCl2 to give ethyl 6-methyl-4-phenyl-2-(oxo/thioxo)-1,2,3,4-tetrahydropyrimidine-5-
carboxylate 1a/1b [5], followed by reaction with hydrazine hydrate in ethanol to give 6-methyl-4-phenyl-2-oxo/
thioxo-1,2,3,4-tetrahydropyrimidine-5-carbohydrazide 2a/2b [5]. Treatment of 6-methyl-4-phenyl-2-oxo/thioxo-
1,2,3,4-tetrahydropyrimidine-5-carbohydrazide 2a/2b with ammonium thiocyanate in acidic medium afforded 6-
methyl-2-(oxo/thioxo)-4-phenyl-1,2,3,4-tetrahydropyrimidine-5-carbonyl hydrazine-carbothioamide, which on het-
erocyclization in presence of cyanogen bromide gave 5-(5-amino-1,3,4-oxadiazol-2-yl)-6-methyl-4-phenyl-3,4-
dihydropyrimidin-2(1H)-one/thione 3a/3b [5,17]. Finally, compounds 3a/3b on condensation with various 5-
substituted indoline-2,3-dione in acidic medium afforded the title compound 5-substituted-3-[{5-(6-methyl-2-oxo/
thioxo-4-phenyl-1,2,3,4 tetrahydropyrimidin-5-yl)-1,3,4-oxadiazol-2-yl}imino]-1,3-dihydro-2H-indol-2-one 4a-l
[5,17]. The structure of all the synthesized compounds was established by elemental, IR, (1H and 13C) NMR and
mass spectral analysis [18].
All the newly synthesized compounds were evaluated in vitro to study its activity against Mycobacterium
tuberculosis H37 Rv. The primary screening was conducted at concentration 6.25 mg/mL in BACTEC MGIT system
[19]. Compounds demonstrating 99% inhibition in the primary screen were described as most potent compounds. The
preliminary results indicated that compounds 4c, 4f, 4i and 4l showed highest inhibition (99%) at a constant
concentration level (6.25 mg/mL). This primary bioassay results have driven us to examine the real potency (MIC) of
the title compounds against M. tuberculosis H37 Rv. The secondary biological screening was performed using
Lowenstein–Jensen MIC method and it is worthwhile to note that compound 4c and 4l displayed inhibition completely
(99%) at the MIC of (3.10–3.12 mg/mL). All the remaining compounds displayed moderate to good activity within
74–99% inhibition at the concentration of (100–500 mg/mL). The secondary antimycobacterial screening for test
compounds was obtained by using L.J. (Lowenstein and Jensen) MIC method [20,21] and the observed MIC of
compounds are presented in Table 1. Isoniazid, Refampin, Ethambutol and Pyrazinamide were used as the reference
drug.
All the synthesized compounds were also evaluated in vitro for antimalarial assay against Plasmodium falciparum
3D7 chloroquine-sensitive strain (Microcare Laboratory & TRC, Surat, Gujarat, India), in 96 well microtitre plates
according to the microassay protocol of Rieckmann and co-workers with minor modifications [22–25]. The test
concentration which inhibited the complete maturation into schizonts was recorded as the minimum inhibitory
concentrations (MIC). Chloroquine was used as the reference drug. Observations of the in vitro antimalarial screening
are presented in Table 1. The compounds 4a, 4b, 4e, 4f, 4j and 4k have no effect on antimalarial activity
(MIC = 10 mg/mL) while compounds 4c, 4d, 4g, 4i, 4j and 4l have remarkable improvement in antimalarial potency
with MIC value in the range of 0.035–5.0 mg/mL. Presence of nitro (4c, MIC = 0.177 mg/mL) and presence of chloro
(4l, MIC = 0.035 mg/mL) displayed excellent antimalarial potency. Overall, among the various substitution, the order

T.N. Akhaja, J.P. Raval / Chinese Chemical Letters 23 (2012) 785–788
787
Table 1
Biological activity of compounds 4a-l.
Entry
Antitubercular activitya
BACTEC MGIT method
Antimalarial activityb
log P valuec
L.J. MIC method
MIC values (mg/mL)
MIC values
%
MIC values
%
(mg/mL)
inhibition
(mg/mL)
inhibition
4a (X = O, R = H)
4b (X = O, R = Br)
4c (X = O,R = NO2)
4d (X = O, R = F)
4e (X = O, R = I)
4f (X = O, R = Cl)
4g (X = S, R = H)
4h (X = S, R = Br)
4i (X = S, R = NO2)
4j (X = S, R = F)
4k (X = S, R = I)
4l (X = S, R = Cl)
Isoniazid
>6.25
>6.25
6.25
–
200
500
3.10
250
500
100
500
500
100
200
500
3.12
0.07
0.25
0.20
–
92
74
99
84
93
97
94
75
97
89
94
99
99
99
99
–
10
10
0.177
5
3.13 Æ 0.83
4.11 Æ 0.88
3.04 Æ 0.84
3.39 Æ 0.88
4.37 Æ 0.88
3.93 Æ 0.84
3.74 Æ 0.85
4.71 Æ 0.89
3.65 Æ 0.86
3.99 Æ 0.89
4.97 Æ 0.89
4.53 Æ 0.86
–
–
99
–
>6.25
>6.25
>6.25
>6.25
>6.25
>6.25
>6.25
>6.25
6.25
–
10
10
5
99
–
–
10
5
99
–
5
–
10
0.035
–
99
99
99
99
99
–
0.20
Refampin
0.25
–
–
Ethambutol
3.12
–
–
Pyrazinamide
6.25
–
–
Chloroquine
–
–
–
0.125
–
a
Against M. tuberculosis H37Rv (MTCC – 200).
b
c
Against Plasmodium falciparum 3D7 chloroquine-sensitive strain.
Theoretical values of log P were calculated using commercially available ChemDraw program.
of highest antimalarial potency is NO2 ꢀ F > Br ꢀ H. It is well known from the literature that the presence of these
groups imparts a variety of properties including steric, electronic properties, enhanced binding interactions, metabolic
stability, changes in physical properties and selective reactivities [26,27]. This promising antitubercular and
antimalarial activity may be due to sufficient hydrogen bonding capacity with desired lipophilicity or with favorable
steric hinderance [28].
Acknowledgment
The author (J.P. Raval) wishes to express his thanks to Chairman – Charutar Vidya Mandal (Dr. C.L. Patel), Director
– SICART (Dr. V.S. Patel) & Director – ARIBAS (Dr. P.S. Patel) for providing necessary research facilities, and Dr.
Dhanji Rajani, Microcare Laboratories, Surat for extending help for assessment of biological activity. We are also
thankful to Mr. Priyakant R. Raval & Dr. Kishor R. Desai, Director, Department of Chemistry, Uka – Tarsadia
University, Bardoli for research advice.
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[18] Analytical data of the synthetic compounds. 3a: Yield 72%, m.p. 210–211 8C, Anal. Calcd. for C13H13N5O2: C, 57.56; H, 4.83; N, 25.82.
Found: C, 57.17; H, 4.48; N, 25.58%. IR ((max, cmÀ1, KBr): 3430, 3067 (C–H, Aromatic), 3384, 3147 (N–H), 1718 (C O), 1634 (C N,
Iminebase), 1458 (C C, Aromatic), 1364 (C N), 1021 (C–O–C), 745, 672, 643 (C–H, Deformation). 1H NMR (400 MHz, DMSO-d6): (8.36,
6.52 (s, 2H, –NH–), 7.42–7.94 (m, 5H, Ar–H), 5.27 (s, 1H, –CH ), 2.37 (s, 3H, –CH3). 13C NMR (100 MHz, DMSO-d6): (161.45 (C O),
154.13, 152.74 (C–O–C), 142.43 (C–CH3), 109.21, 127.28, 132.81, 136.82 (Ar–C), 111.17 (C–oxadiazole ring), 54.25 (C–furfural ring), 13.98
(–CH3). MS [M]+ [272.36]+. 3b: Yield 79%, m.p. 222–223 8C, Anal. Calcd. for C13H13N5OS: C, 54.34; H, 4.56; N, 24.37. Found: C, 53.89; H,
3.93; N, 23.76%. IR ((max, cmÀ1, KBr): 3430, 3067 (C–H, Aromatic), 3371, 3154 (N–H), 1624 (C N, Iminebase), 1462 (C C, Aromatic),
1429 (C S), 1372 (C N), 1027 (C–O–C), 736, 693, 632 (C–H, Deformation). 1H NMR (400 MHz, DMSO-d6): (8.36 (s, 2H, –NH–), 7.12–
7.57 (m, 5H, Ar–H), 5.46 (s, 1H, –CH ), 2.18 (s, 3H, –CH3). 13C NMR (100 MHz, DMSO-d6): (173.17 (C S), 155.23, 153.87 (C–O–C),
142.68 (C–CH3), 110.29,125.27, 132.45, 139.13 (Ar–C), 113.37 (C–oxadiazole ring), 55.76 (C–furfural ring), 14.39 (–CH3). MS [M]+
[288.57]+. 4c: Yield 90%, m.p. 251–253 8C, Anal. Calcd. for (C21H15N7O5): C, 56.63; H, 3.39; N, 17.96. Found: C, 55.93; H, 3.05; N, 17.61%.
IR ((max, cmÀ1, KBr): 3430, 3067 (C–H, Aromatic), 3371, 3189 (N–H), 1712 (C O), 1631 (C N, Iminebase), 1565 (N O), 1459 (C C,
Aromatic), 1375 (C N), 1252, 1024 (C–O–C), 741, 685, 657 (C–H, Deformation). 1H NMR (400 MHz, DMSO-d6): (9.05, 6.36 (s, 3H, –NH–
), 7.03–7.69 (m, 8H, Ar–H), 5.23 (s, 1H, –CH ), 2.15 (s, 3H, –CH3). 13C NMR (100 MHz, DMSO-d6): (171.29, 168.34 (C O), 155.45
(C N–), 153.72, 151.65 (C–O–C), 97.12, 104.37, 110.53, 114.27, 121.37, 126.38, 133.54, 135.65, 142.45, 149.32 (Ar–C), 57.43 (C–furfural),
14.78 (–CH3). MS [M]+ [447.17]+. 4l: Yield 79%, m.p. 245–246 8C, Anal. Calcd. for (C21H15ClN6O2S): C, 55.94; H, 3.35; N, 18.64. Found: C,
55.71; H, 3.12; N, 18.47%. IR ((max, cmÀ1, KBr): 3437, 3043 (C–H, Aromatic), 3365, 3183 (N–H), 1718 (C O), 1643 (C N, Iminebase),
1457 (C C, Aromatic), 1367 (C N), 1246, 1029 (C–O–C), 746, 667, 632 (C–H, Deformation). 710 (C–Cl). 1H NMR (400 MHz, DMSO-d6):
d 11.92, 9.34, 2.67 (s, 3H, –NH–), 7.18–7.93 (m, 8H, Ar–H), 5.12 (s, 1H, –CH ), 2.29 (s, 3H, –CH3). 13C NMR (100 MHz, DMSO-d6): d
173.67 (C S), 166.75 (C O), 155.65 (C N–), 153.68, 152.05 (C–O–C), 97.12, 107.45, 110.32, 115.53, 121.29, 126.54, 132.48, 136.84,
143.23, 149.19 (Ar–C), 56.96 (C–furfural), 16.19 (–CH3). MS [M]+ [452.07]+.
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