Synthesis and antituberculosis activity of some new 2-quinoxalinecarbonitriles.

Article abstract of DOI:10.1016/S0014-827X(98)00067-6

Tuberculosis, an ancient disease undergoing recent control by public hygiene and drug therapy, has experienced a recrudescence throughout the world. New and effective therapies are rapidly needed to combat infections caused by these strains. Some new 2-quinoxalinecarbonitriles have been synthesized and tested as antituberculosis agents and interesting results have been obtained from the first screening.

Full text of DOI:10.1016/S0014-827X(98)00067-6

Il Farmaco 53 (1998) 570±573
Synthesis and antituberculosis activity of some
new 2-quinoxalinecarbonitriles
a
M. Elena Montoya , Yolanda Sainz , M. Angel Ortega , Adela Lopez De Cerain ,
b
b
b
b,
*
Antonio Monge
a
Facultad de Farmacia y Bioqu Âõ mica, Universidad Nacional Mayor de San Marcos, Lima, Peru
Centro de Investigaci o n en Farmacobiolog Âõ a Aplicada, Departamento de Qu Âõ mica Org a nica y Farmac e utica, Facultad de Farmacia,
Universidad de Navarra, 31080 Pamplona, Spain
b
Received 10 March 1998; accepted 8 September 1998
Abstract
Tuberculosis, an ancient disease undergoing recent control by public hygiene and drug therapy, has experienced a recrudescence
throughout the world. New and effective therapies are rapidly needed to combat infections caused by these strains. Some new 2-quinox-
alinecarbonitriles have been synthesized and tested as antituberculosis agents and interesting results have been obtained from the ®rst
screening. q 1998 Elsevier Science S.A. All rights reserved.
Keywords: Antituberculosis activity; 2-Quinoxalinecarbonitriles; Tuberculosis
1
. Introduction
and some quinoxaline 1,4-di-N-oxide derivatives with very
different substituents in the 2, 3, 6 and 7 positions have
shown antituberculosis activity [10,11].
Mycobacterium tuberculosis infects over one-third of the
world's population and causes almost three million deaths
every year [1±3]. The World Health Organization suggests
that at least 5.6 million persons worldwide, mostly in devel-
oping countries, carry dual infection, and . 1.4 million
cases of HIV-related tuberculosis will occur by the year
We described substituted quinoxaline 1,4-di-N-oxides.
This type of structure seemed fundamental for the intended
activity. Now, we propose to establish the importance of
quinoxaline1,4-di-N-oxides derivatives versus quinoxaline
derivatives.
2000 [4]. Ethambutol is one of the primary drugs used in
combination with isoniazid, rifampin, streptomycin and
pyrazinamide to treat tuberculosis. But M. tuberculosis is
often resistant to multiple drugs, therefore antituberculosis
agents with bactericidal mechanisms different from those of
available ®rst-line drugs, are urgently needed [5].
2
. Chemistry
-Quinoxalinecarbonitriles derivatives were prepared
2
using the synthetic methods outlined in Scheme 1. The start-
ing products 5(6)-substituted benzofuroxane (1a±1c), and 3-
amino-6(7)-substituted-2-quinoxalinecarbonitrile 1,4-di-N-
oxides (2a±2c), were obtained by previously described
methods [12]. Compounds 2a, 2b or 2c were reacted with
sodium ditionite in methanol to give 3-amino-6(7)-substi-
tuted-2-quinoxalinecarbonitriles (3a±3c); 3a had been
described previously [13] and 3b, 3c were prepared from
a similar synthesis, but the puri®cation process was different
to that described. We obtained isomer mixtures in both cases
although in different proportions. The reaction of 3a, 3b or
Some quinoxalines and quinoxalines 1,4-di-N-oxides
were synthesized and evaluated as antibacterial agents [6±
8]. Other quinoxaline 1,4-di-N-oxides were similar to vita-
min K, and their use for the treatment of the tuberculosis
was suspected in the past [9]. The possibility that these type
of compounds could be useful for treatment of tuberculosis
was proposed. With this idea, some new quinoxaline 1,4-di-
N-oxides were prepared in our laboratory and screened at
the GWL Hansen's Disease Center within the TAACF
(Tuberculosis Antimicrobial Acquisition Coordinating
Facility). The primary screening has been very interesting
3
6
c with a fresh solution of sodium nitrite gave 3-chloro-
(7)-substituted-2-quinoxalinecarbonitriles (4a±4c); 4a
had been described previously [14] and we prepared 4b,
4c from a similar synthesis, but the puri®cation of these
*
Corresponding author.
0014-827X/98/$ - see front matter q 1998 Elsevier Science S.A. All rights reserved.
PII S0014-827X(98)00067-6

M.E. Montoya et al. / Il Farmaco 53 (1998) 570±573
571
Table 2
Elemental analysis
Compound
C
H
N
3
3
4
4
4
b
c
Calc.
Found
Calc.
52.82
52.88
50.43
50.04
56.99
57.38
48.24
48.58
46.62
46.89
2.46
2.43
2.11
2.13
2.11
2.24
1.34
1.38
1.17
1.15
27.38
27.54
23.52
23.25
22.16
21.67
18.75
18.77
16.31
16.34
Found
Calc.
a
b
c
Found
Calc.
Found
Calc.
Found
to inhibit the growth of virulent Mycobacterium tuberculo-
sis.
4
. Results and discussion
Scheme 1.
The results of in vitro evaluation of antituberculosis activ-
products, 4a±4c, was different to that described; 4a and 4c
were obtained as isomer mixtures.
All the compounds were characterized by physical
constants (Table 1), elemental analysis (Table 2), IR, H
NMR and MS spectra.
ity appears in Table 3. The minimum inhibitory concentra-
tion (MIC) is . 12.5 mg/ml for 3b, 3c, 4b and 4c, and is
, 12.5 mg/ml for 4a. The last compound, which shows
an inhibition greater than 90% (94), has been selected
for con®rmatory and advanced screening. The other
compounds, considered inactive, still have signi®cant inhi-
bitory activity (inhibition from 23 to 68).
1
We can establish preliminary structure±activity relation-
ships on the basis of the primary screening. If we compare
these results with those obtained for quinoxaline 1,4-di-N-
oxides [10,11], it seems that the N-oxide function is impor-
tant for the activity: seven 3-amino or 3-amino-substituted
3
. Pharmacology
Compounds 3b, 3c, 4a, 4b and 4c were tested for their
antituberculosis activity. In vitro evaluation of antitubercu-
losis activity was carried out at the GWL Hansen's Disease
Center (Colorado State University) within the TAACF
2
-quinoxalinecarbonitrile 1,4-di-N-oxides with one or two
chloros in position 6(7) are active [10,11], and the
compound 3b, with the same structure but not N-oxide,
does not present activity. But according to these results,
the presence of the N-oxide moiety in the structure is not
fundamental; the compound 4a, 3-chloro-2-quinoxalinecar-
bonitrile, shows an inhibition of 94% (MIC , 12.5 mg/ml).
In the case of the inactive compounds, the 3-chloro substi-
tution with the 6(7)-chloro derivative is better than the 3-
amino substitution (4b versus 3b, with 58 and 23% inhibi-
tion respectively), and the 3-amino substitution with the
(
Tuberculosis Antimicrobial Acquisition Coordinating
Facility) screening program for the discovery of novel
drugs for treatment of tuberculosis. The purpose of the
screening program is to provide a resource whereby new
experimental compounds can be tested for their capacity
Table 1
Physicochemical properties of the tested compounds
6
(7)-tri¯uoromethyl derivative is better than the 3-chloro
Table 3
In vitro evaluation of antituberculosis activity: rifampin (RMP) MIC 0.25
mg/ml versus Mycobacterium tuberculosis
Compound
R
3
R
7
Formula
M.p. (8C)
Yield (%)
Compound
MIC (mg/ml)
% Inhibition
3
3
4
4
4
b
NH
NH
Cl
2
Cl
CF
H
C
C
C
C
C
9
H
5
ClN
4
245 (d)
198±201
156±158
140±142
45±47
75
65
42
46
42
a
a
c
2
3
10
9
H
5 3 4
F N
3
3
b
c
. 12.5
. 12.5
, 12.5
. 12.5
. 12.5
23
68
94
58
37
a
b
c
H
4
ClN
ClF
Cl
3
Cl
Cl
CF
10
H
3
3 3
N
4a
Cl
3
9
H
3
2
N
3
4b
a
4c
6 7
Isomeric mixture (R or R ).

5
72
M.E. Montoya et al. / Il Farmaco 53 (1998) 570±573
substitution (3c versus 4c, with 68 and 37% inhibition
respectively).
5.1.1.2. 3-Amino-6(7)-tri¯uoromethyl-2-quinoxaline-
carbonitrile (3c)
On the basis of these results, 3-chloro-2-quinoxalinecar-
bonitrile shows good antituberculosis activity in the primary
screening. We propose to con®rm the results, to advance
further on in the screening, to synthesize new 2-quinoxali-
necarbonitrile compounds with different substituents in
positions 3, 6 and 7 in order to get the best antituberculosis
activity, and to study these structures beside the correspond-
ing N-oxides, because quinoxalines versus quinoxaline 1,4-
di-N-oxides may be better from the point of view of biolo-
gical biodisponibility.
The mixture reaction was stirred for 3 h. The product was
puri®ed by recrystallization from water/dimethylformamide
to afford 3c (65% yield) as yellow solid, m.p. 198±2018C.
An isomeric mixture, 6 and 7-tri¯uoromethyl, was obtained
2
1
(50:50). IR (KBr, cm ) n 1172 (C±F), 1560 (CyN), 1664
1 200
(NH ), 2236 (CxN), 3331 (NH); H NMR [(CD ) SO,
2
3 2
ppm] d 7.67±7.76 (m, 2H, H (7-tri¯uomethyl) 1 H (6-
6
7
tri¯uoromethyl)), 7.86 (s, 2H, NH (7-tri¯uoromethyl)),
2
7.87 (s, 1H, H (6-tri¯uoromethyl)), 7.96 (dd, H, H (7-
5
5
tri¯uoromethyl), J5±7 ˆ 1:8 Hz, J ˆ 8:9 Hz), 8.06 (d,
7±8
1
H, H (6-tri¯uoromethyl), J ˆ 8:7 Hz), 8.22 (s,1H, H
8
7±8
8
1
7-tri¯uoromethyl)); MS: m/z (%) 238 (M , 100), 211 (36),
z
(
159 (16), 132 (7), 117 (4), 75 (39); Anal. C H F N (C, H,
N).
5. Experimental
1
0
5
3
4
5.1. Chemistry
5.1.2. General procedure for the preparation of 3-chloro-2-
quinoxalinecarbonitrile derivatives (4a±4c)
IR spectra were recorded on a Perkin-Elmer 681 infrared
1
spectrophotometer, using KBr pellets. H NMR spectra
A mixture of 6(7)-substituted-3-amino-2-quinoxalincar-
bonitrile (5 mmol), acetic acid (18 ml) and hydrochloric
acid 35% (22 ml) was stirred at 08C. A fresh solution of
sodium nitrite (20 mmol) was added. The reaction mixture
was stirred for some hours at room temperature. The result-
ing precipitate was collected and washed with cool water.
The impure product was puri®ed by ¯ash chromatography
(
(
reference TMS int) were taken on a Bruker AC-200E
200 MHz). The mass spectra were recorded on a
Hewlett-Packard 5988-A instrument at 70 eV. Column
chromatography was performed on a Merck silica gel 200
ASTM and analytical TLC on Panreac Silica gel 0.2 mm
aluminium sheets. The plates were scanned under ultraviolet
light at 254 and 366 nm. Melting points were determined on
a Mettler FP82 1 FP80 apparatus and are uncorrected.
Elemental analysis were obtained on an Elemental Analyzer
from vacuum-dried samples (over P O at 1±2 mm Hg, 24 h
(
SP, silica gel; eluting with n-hexane/ethyl acetate).
5
.1.2.1. 3-Chloro-2-quinoxalinecarbonitrile (4a)
The mixture reaction was stirred for 1 h. The product was
2
5
at 60±808C) and were within 0.4 % of the theoretical values.
Compounds 1a±1c, 2a±2c, 3a and 4a have been reported
previously [12±14].
puri®ed by ¯ash chromatography, eluting with n-hexane/
ethyl acetate (75:25) to afford 4a (42% yield) as white
21
solid, m.p. 156±1588C. IR (KBr, cm ) n 1556 (CyN),
1
200
2
[
234 (CxN), 3039 (C±H aromatic);
(CD ) SO, ppm] d 8.03±8.13 (m, 3H, H 1 H6 1 H ),
H
NMR
5.1.1. General procedure for the preparation of 3-amino-2-
quinoxalinecarbonitrile derivatives (3b, 3c)
3
2
5
7
1
z
8
1
,24 (d, 1H, H J
8
ˆ 8:4 Hz); MS: m/z (%) 189 (M ,
ꢀ7±8†
00), 154 (33), 102 (32), 76 (19). Anal. C H ClN (C, H, N).
9 4 3
,
A solution of 6(7)-substituted-2-quinoxalinecarbonitrile
,4-di-N-oxide (5 mmol) in methanol (20 ml) was stirred
1
at 508C. A fresh solution of sodium ditionite (20 mmol) was
added. The reaction mixture was allowed to cool, and the
resulting crystals were collected and washed with water.
5.1.2.2. 3,7-Dichloro-2-quinoxalinecarbonitrile (4b)
The reaction mixture was stirred for 1 h. The product was
puri®ed by ¯ash chromatography, eluting with n-hexane/
ethyl acetate (90:10) to afford 4b (46% yield) as white
2
1
5
.1.1.1. 3-Amino-6(7)-chloro-2-quinoxalinecarbonitrile
3b)
The mixture reaction was stirred for 1 h. The product was
solid, m.p. 140±1428C. IR (KBr, cm ) n 1547 (CyN),
1
200
(
2237 (CxN), 3041 (C±H aromatic);
[(CD ) SO, ppm] d 8.06±8.27 (m, 2H, H 1 H ), 8.43 (s,
H
NMR
3
2
5
6
1z
puri®ed by recrystallization from methanol/dimethyl-
formamide to afford 3b (75% yield) as yellow solid, m.p.
1H, H ); MS: m/z (%) 223 (M , 100), 136 (24), 110 (18), 75
8
(20); Anal. C H Cl N (C, H, N).
3
9
3
2
2
major isomer was the 7-chloro isomer. IR (KBr, cm ) n
458C (d). An isomeric mixture was obtained, in which the
2
1
5.1.2.3. 3-Chloro-6(7)-tri¯uoromethyl-2-quinoxaline-
carbonitrile (4c)
1
1
NMR
560 (CyN), 1656 (NH ), 2232 (CxN), 3316 (NH); H
2
200
[(CD ) SO, ppm] d 7.44±7.60 (m, 3H, H (7-
3
The reaction mixture was stirred for 2 h. The product was
puri®ed by ¯ash chromatography, eluting with n-hexane/
ethyl acetate (85:15) to afford 4c (42% yield) as white solid,
m.p. 45±478C. An isomeric mixture of 6- and 7-
2
6
chloro) 1 H (7-chloro) 1 H (6-chloro)), 7.56 (s, 2H,
7
5
NH ), 7.73 (d, 1H, H (6-chloro), J ˆ 8:9 Hz), 7,85 (d,
2
5
5±6
1
H (6-chloro), J ˆ 8:8 Hz), 7.92 (s,1H, H , (7-chloro));
8
7±8
8
1
z
21
MS: m/z (%) 204 (M , 100), 177 (32), 125 (10), 110 (5), 75
9); Anal. C H ClN , (C, H, N).
tri¯uoromethyl was obtained (50:50). IR (KBr, cm ) n
1130 (C±F), 1560 (CyN), 2236 (CxN), 3073 (C±H
(
9
5
4

M.E. Montoya et al. / Il Farmaco 53 (1998) 570±573
573
1 200
aromatic); H NMR [(CD ) SO, ppm] d 8.38±8.29 (m,
tuto de Cooperaci o n Iberoamericana (I.C.I.) for a predoc-
toral grant to M. Elena Montoya and to Gobierno de Navarra
for a predoctoral grant to M. Angel Ortega.
3
2
4
H, H (7-tri¯uoromethyl) 1 H (7-tri¯uoromethyl) 1 H
5 6
7
(
8
(
6-tri¯uoromethyl)), 8.49 (d, H (6-tri¯uoromethyl), J7±8
8
ˆ
:7 Hz), 8.63 (s, 1H, H (6-tri¯uoromethyl)), 8.74 (s, 1H, H
5
8
1
7-tri¯uoromethyl)); MS: m/z (%) 257 (M , 79), 222 (26),
z
1
70 (62), 144 (28), 75 (100); Anal. C H ClF N (C, H, N)
10 3 3 3
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5.2. Pharmacology
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