Synthesis, characterization of novel coupling products and 4-arylhydrazono-2-pyrazoline-5-ones as potential antimycobacterial agents

Article abstract of DOI:10.1016/S0014-827X(02)01253-3

Novel coupling products 7a-d and 4-arylhydrazono-2-pyrazoline-5-ones 8a-e were synthesized and evaluated for antimycobacterial activity against Mycobacterium tuberculosis H37Rv and Mycobacterium avium. Compound 7b was found to be the most potent derivatives of the 7a-d series by an MIC value of 6.25 μg/ml.

Full text of DOI:10.1016/S0014-827X(02)01253-3

Il Farmaco 57 (2002) 583–587
www.elsevier.com/locate/farmac
Synthesis, characterization of novel coupling products and
-arylhydrazono-2-pyrazoline-5-ones as potential antimycobacterial
agents
4
S¸ . G u¨ niz K u¨ c¸ u¨ kg u¨ zel, Sevim Rollas*
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Marmara Uni6ersity, Tibbiye cad. No. 49, Haydarpa s¸ a 81010 I
:
stanbul, Turkey
Received 26 April 2002
Abstract
Novel coupling products 7a–d and 4-arylhydrazono-2-pyrazoline-5-ones 8a–e were synthesized and evaluated for antimycobac-
terial activity against Mycobacterium tuberculosis H37Rv and Mycobacterium a6ium. Compound 7b was found to be the most
´
potent derivatives of the 7a–d series by an MIC value of 6.25 mg/ml. © 2002 Published by Editions scientifiques et m e´ dicales
Elsevier SAS.
Keywords: Coupling products; 4-Arylhydrazono-2-pyrazoline-5-one; 1,2,4-Triazole; Hydrazide–hydrazones
1. Introduction
2. Chemistry
Isoniazide and hydrazones of isoniazide [1–5], some
-aminobenzoic acid hydrazide–hydrazones [6] and
their coupling products with indole [7], 1,3,4-oxadiazo-
line-2(3H)-thione starting from p-aminosalicylic acid
hydrazide [8], pyrazolone derivatives [9], 1-methyl-1H-
During the course of this study, some novel coupling
products containing hydrazide–hydrazone moiety; 2-
pyrazoline-5-one derivatives obtained from either 1,3,4-
oxadiazoline-2(3H)-thione or 1,2,4-triazoline-3(2H)-
4
2-imidazo [4][5b] pyridine carboxylic acid hydrazide–
hydrazones [10], 1,2,4-triazole derivatives [11] have
been reported to possess promising antitubercular acti-
vity against Mycobacterium tuberculosis H37Rv. In our
earlier studies, coupling products of 5-nitro-2-furylhy-
drazone (A) [12], 4-arylhydrazono-2-pyrazoline-5-ones
from 1,3,4-oxadiazole-2(3H)-thione derivatives (B) [13],
1,2,4-triazoline-3(2H)-thione derivatives containing
thiourea moiety (C) [11] examined by the Tuberculosis
Antimicrobial Acquisition & Coordinating Facility
(
TAACF), have been proved to possess considerable
inhibition against M. tuberculosis H37Rv (Fig. 1).
Therefore, it was planned to design more potent an-
timycobacterial agents, effective in lower dose accom-
panying with low toxicity.
*
Corresponding author
Fig. 1. Some compounds proved to inhibit the growth of M. tubercu-
losis H37R6.
E-mail address: sevim@sevimrollas.com (S. Rollas).
´
0
014-827X/02/$ - see front matter © 2002 Published by Editions scientifiques et m e´ dicales Elsevier SAS.
PII: S0014-827X(02)01253-3

5
84
S¸ .G. K u¨ c¸ u¨ kg u¨ zel, S. Rollas / Il Farmaco 57 (2002) 583–587
Fig. 2. Preparation of the aromatic primary amines 3, 5 and 6. Key to reactions: (i) C H COCl–ether; (ii) H NꢀNH ·H O–EtOH-reflux; (iii)
6
5
2
2
2
CH NCS–EtOH-reflux; (iv) (1) NaOH(2N)-reflux, (2) HCl; (v) 5-nitrofurfural–EtOH-reflux; (vi) (1) CS –KOH-reflux, (2)HCl.
3
2
thiones have been synthesized (Figs. 2 and 3). Purity of
these compounds has been confirmed by elemental
analysis whilst their structures were elucidated using
22]. EI-MS spectra of 7a, 7b, 7c and 7d confirmed their
molecular weights displaying the parent molecular ions
at m/z 389, 415, 447 and 347, respectively.
1
UV and H NMR spectroscopy. Antituberculosis pro-
perties of the new compounds, synthesized in the
present study, have been investigated in TAACF.
The UV spectra of 7a–d and 8a–e exhibited charac-
teristic K bands for chromophoric CꢁN group at 365–
3. Experimental
3.1. Chemistry
385 and 395–433 nm regions, respectively, which
corresponds to hydrazone groups. In the UV spectra of
compounds 7a–d and 8a–e, absorptions arising from
Benzocaine, carbondisulfide and 4-nitro-phenylhy-
drazine were purchased from Merck. All other chemi-
cals were purchased from Fluka. All melting points
NꢁN bond at around 332–360 nm [14,15] and above
1
4
00 nm [16] were not observed. The H NMR spectra
(
m.p.) were recorded on a Buchi-530 m.p. apparatus
of 7a–c and 8a–c, 8e in DMSO-d displayed the hydra-
6
zone NꢀH protons at 11.92–13.56 ppm. The hydrazone
and triazoline protons of 7d and oxadiazoline protons
of 8d were observed to exchange with deuterium in
1
DMSO-d . In addition, H NMR spectra of these com-
6
pounds signals arising from possible ꢂCHꢀNꢁNꢀ
structure at 3.00–4.00 ppm [17,18] were not observed.
1
UV and H NMR findings of these compounds also
supported that the structures of 7a–d and 8a–e might
be given in hydrazone form. The azomethine (CHꢁN)
and NꢀH protons (CONHNꢁCH) of hydrazide–hydra-
zone 7a–c exhibited the singlets at 8.37–8.44 ppm. The
NꢀH protons of triazoline (8a–c) and oxadiazoline (8e)
resonated at 13.93, 13.75, 13.93 and 14.30–15.23 ppm.
1
H NMR spectrum of 8d has been taken by heating due
1
to low solubility of this compound. For this reason, H
NMR spectrum of 8d has been displayed shift to low
energy medium. The carboxylic acid protons of com-
pound 7a were observed to exchange with deuterium in
Fig. 3. Synthesis of target compounds 7a–e and 8a–e (*Reference
26]) Key to reactions: (i) NaNO +2HCl/0–5 °C; (ii)
[
2
R ꢀCOꢀCH ꢀCOꢀR –AcONa; (iii) R ꢀC H ꢀNHꢀNH –AcOH-
1
2
2
3
6
4
2
DMSO-d , which was supported by the literature [19–
reflux.
6

S¸ .G. K u¨ c¸ u¨ kg u¨ zel, S. Rollas / Il Farmaco 57 (2002) 583–587
Molecular weight EI-MS (m/z, 70 eV)
585
Table 1
Physical and spectral data of 7a–d
Comp. M.p. (°C)
Yield (%)
UV umax log m (1
(
M.Wt.)
mg per 100 ml)
+
7
7
7
7
a
b
c
260–270
decomposition)
234–240
decomposition)
245–260
decomposition)
165–170
63
71
68
80
C15H11N O
(389)
389 [M ], 254, 235, 208, 207, 164, 154, 131, 121, 119,
104,93
380 (4.37)
302 (4.17)
381 (4.53)
302 (4.17)
206 (4.58)
5
8
7
6
(
+
C18H17N O
415 [M ], 274, 273, 254, 120, 104, 92
5
(
(415)
+
C22H17N O
447 [M ], 331, 309, 308, 293, 274, 273, 251, 228, 224, 223,
5
(
(447)
147, 119
+
d
C15H17N O S
347 [M ], 315, 302, 275, 274, 232, 207, 206, 205, 191, 190,
365 (4.43)
256 (4.33)
5
3
(
347)
176, 133, 132, 118
Table 2
Physical, analytical and spectral data of 8a–e
Compd.
M.p. (°C)
Yield (%)
Molecular weight (M.Wt.)
Elemental analysis
UV umax log m (1 mg per 100 ml)
C
H
S
8
a
b
285–287
287–289
70
63
C19H17N OS·1/2H O (400.46)
56.99
4.53
4.46
8.00
7.47
401 (4.34)
245 (4.70)
7
2
5
7.04
2
10 (4.86)
8
C19H16N O S (436.46)
52.28
1.89
3.69
3.23
7.34
7.00
395 (3.71)
235 (4.54)
8
3
5
2
1
09 (4.82)
94 (4.10)
8
c
280–282
\300
67
56
C20H19N O S·1/2H O (430.48)
55.80
5.88
50.00
0.94
4.68
4.53
3.26
3.10
7.45
7.39
7.41
7.14
399 (3.68)
209 (4.78)
433 (3.70)
301 (3.69)
7
2
2
5
8
d
C18H13N O S·1/2H O (432.41)
7 4 2
5
2
2
1
34 (4.56)
09 (4.82)
92 (4.08)
8
e
287–290
58
C19H16N O S·1/2H O (417.44)
55.87
5.62
3.95
3.85
7.85
7.57
428 (4.47)
295 (4.23)
6
3
2
5
2
2
44 (4.69)
09 (4.86)
and uncorrected. UV spectra were recorded on a Shi-
madzu UV 2100S spectrophotometer (1 mg per 100 ml
in ethanol). Nuclear Magnetic Resonance spectra ( H
NMR), were recorded on a Bruker AVANC-DPX 400
spectrometer. MS spectra were obtained on a Fisons
Instruments VG Platform II LC-MS in the electron
impact (EI) mode. Elemental analyses were performed
on a Carlo Erba 1106 instrument.
3.1.3. Synthesis of 4-arylhydrazono-2-pyrazoline-5-ones
(8a–e)
1
The coupling products 7d and 7e were refluxed with
phenylhydrazine–4 - nitrophenylhydrazine–4 - methoxy-
phenylhydrazine hydrochloride to yield 8a–e using
a reported procedure [13,27,28]. Physical and spec-
tral data of these compounds are given in Tables 2
and 3.
3.1.1. Preparation of aromatic primary amines (3, 5, 6)
3.2. Biological acti6ity
These compounds were prepared as described pre-
viously [23–25].
Compounds were also tested for in vitro antimy-
cobacterial activity against M. tuberculosis H37Rv
using the BACTEC 460 radiometric system [29].
The effective compounds were also tested against My-
cobacterium a6ium using the same technique. Inhibi-
tions of the synthesized compounds against M.
tuberculosis H37Rv and M. a6ium are given in Tables 4
and 5.
3.1.2. Synthesis of coupling products (7a–e)
Compounds 7a–e were synthesized by the reactions
of diazonium salts of 3,5,6 with malonic acid–ethyl
acetoacetate–benzoylacetone according to literature
methods [12,26]. Physical and spectral data of these
compounds are given in Tables 1 and 2.

5
86
S¸ .G. K u¨ c¸ u¨ kg u¨ zel, S. Rollas / Il Farmaco 57 (2002) 583–587
Table 3
1
H NMR spectral data of 7a–d and 8a–e
Compd.
1H NMR l (ppm)
7
7
a
b
7.19–7.79 (m, 6H, ArꢀH); 8.37 (s, 1H, CHꢁN); 11.83 (s, 1H, CONHNꢁCH); 11.92 (b, 1/2H, NHꢀNꢁCꢃ)
1.29 (t, 3H, CH ꢀCH ); 2.42 (s, 3H, COꢀCH ); 4.32 (q, 2H, CH ꢀCH ); 6.61–7.79 (m, 6H, ArꢀH); 8.37 (s, 1H, CHꢁN); 11.93
2
3
3
2
3
(
s, 1H, CONHNꢁCH); 12.29 (b, 1/2H, NHꢀNꢁCꢃ)
7
7
8
8
8
c
d
a
b
c
2.08 (s, 3H, COꢀCH ); 7.27–8.31 (m, 12H, ArꢀH); 8.44 (s, 1H, CHꢁN); 12.21 (s, 1H, CONHNꢁCH); 12.97 (b, 1/2H,
NHꢀNꢁCꢃ)
3
1.26 (t, 3H, CH ꢀCH ); 2.36 (s, 3H, COꢀCH ); 3.54 (s, 3H, NꢀCH ); 4.24 (q, 2H, CH ꢀCH ); 7.50 (d, 2H, o-NH, J=8.5
2
3
3
3
2
3
Hz); 7.65 (d, 2H, m-NH, J=8.5 Hz)
2.32 (s, 3H, pyrazoline CH ); 3.56 (s, 3H, triazoline NꢀCH ); 7.21–7.93 (m, 9H, ArꢀH); 12.99–13.49 (s, 1H, hydrazone NꢀH);
3
3
1
3.93 (s, 1H, triazoline NꢀH)
2.21 (s, 3H, pyrazoline CH ); 3.41 (s, 3H, triazoline NꢀCH ); 7.67–8.22 (m, 4H, ArꢀH); 12.50–13.50 (b, 1H, hydrazone NꢀH);
3
3
1
3.75 (s, 1H, triazoline NꢀH)
2.30 (s, 3H, pyrazoline CH ); 3.56 (s, 3H, triazoline NꢀCH ); 3.77 (s, 3H, OꢀCH ); 7.01–7.82 (m, 8H, ArꢀH); 12.80–13.56 (b,
3
3
3
1
H, hydrazone NꢀH); 13.93 (s, 1H, triazoline NꢀH)
8
8
d
e
4.87 (s, 3H, pyrazoline CH ); 10.27–10.84 (m, 8H, ArꢀH)
3
2.31 (s, 3H, pyrazoline CH ); 3.78 (s, 3H, OꢀCH ); 7.02–7.94 (m, 8H, ArꢀH); 12.56–13.74 (b, 1H, hydrazone NꢀH);
3
3
1
4.30–15.23 (b, 1H, oxadiazoline NꢀH)
4
. Results and discussion
not evaluated further. Compounds 7b and 7c effecting
90% inhibition in the primary screen at 12.5 mg/ml
\
The synthesized compounds in the present study were
was re-tested at lower concentrations against M. tuber-
culosis H37Rv. Level II assay results of these com-
pounds are given in the Table 4. Compound 7b which
was obtained from coupled diazonium salt of 4-
aminobenzoic acid [(5-nitro-2-furyl)methylene]hydra-
zide with benzoylacetone was found to be active against
M. tuberculosis H37Rv at 6.25 mg/ml. The same com-
pounds were also tested against M. a6ium using the
same technique. Clarithromycin was used as the stan-
dard in the tests. No satisfactory level of inhibition was
observed with compounds 7b and 7c at 12.5 mg/ml,
whereas clarithromycin exhibited 98% inhibition at 2
mg/ml. However, in our previous study [12], 4-
aminobenzoic acid [(5-nitro-2-furyl)methylene]hydra-
zide 3, which was starting compound of 7b and 7c, was
observed to inhibit the growth of M. tuberculosis
H37Rv by only 70%. From the given data, it can be
concluded that 7b could be a leading compound for
further development.
tested for their antimycobacterial activity against M.
tuberculosis H37Rv and M. a6ium using the BACTEC
460 radiometric system [29]. The results are shown in
Tables 4 and 5. Rifampicin was used as the standard in
the assays for M. tuberculosis H37Rv. The compounds
which exhibited B90% inhibition in the primary screen
(
Initial screening concentrations were 12.5 mg/ml for 7b
and 7c, 6.25 mg/ml for the remaining compounds) were
Table 4
Antimycobacterial results of 7a–d and 8a–e against M. tuberculosis
H37Rv
Compd.
MIC (mg/ml)
Inhibition (%)
Level II
7
7
7
7
8
8
8
8
8
a
b
c
d
a
b
c
d
e
\6.25
B12.5
B12.5
\6.25
\6.25
\6.25
\6.25
\6.25
\6.25
0.25
0
94
95
1
0
1
1
0
2
98
6.25
12.5
Acknowledgements
Rifampicin
This work was supported by the Research Fund of
Marmara University Project number: SBDYD 130-131-
1
49. We thank Dr. Joseph A. Maddry from the Tuber-
Table 5
culosis Antimicrobial Acquisition and Coordinating
Facility (TAACF), National Institute of Allergy and
Infections Diseases Southern Research Institute, GWL
Hansen’s Disease Center Colorado State University,
Birmingham, AL, USA, for the in vitro evaluation of
antimycobacterial activity using M. tuberculosis H37R6
and M. a6ium.
Antimycobacterial results of 7b and 7c against M. a6ium
Compd.
MIC (mg/ml)
Inhibition (%)
7
7
b
c
\12.5
\12.5
2
45
16
98
Clarithromycin

S¸ .G. K u¨ c¸ u¨ kg u¨ zel, S. Rollas / Il Farmaco 57 (2002) 583–587
587
[
15] H.G. Garg, V. Arora, Chemistry and biological activity of
N%-acyl-4-arylazopyrazoles, J. Pharm. Sci. 61 (1971) 130–132.
16] J. Elguero, R. Jacquier, G. Tarrago, Structure des produits de
copulation du chlorure de ph e´ nyldiazonium avec les ß-dic e´ ke-
tones et les ß-c e´ toesters, Bull. Soc. Chim. 9 (1966) 2981–2989.
17] N. Ergen c¸ , S. Rollas, The coupling products of aliphatic active
C-H compounds with diazonium salts, J. Fac. Pharm. Istanbul.
References
[
[
[
[
[
[
1] E.M. Bavin, D.J. Drain, M. Seiler, D.E. Seymour, Some further
studies on tuberculostatic compounds, J. Pharm. Pharmacol. 4
(1952) 844–855.
[
[
[
[
[
2] N.G. Buu-Hoi, D. Xuong, H. Nam, F. Binon, R. Royer, Tuber-
culostatic hydrazides and their derivatives, J. Chem. Soc. (1953)
1
1 (1975) 8–23.
1
358–1364.
18] N. Ergen c¸ , S. Rollas, p-(Acetylacetonylidenhydrazino)-
sulfaguanidine and two new azopyrazoles, J. Fac. Pharm. Istan-
bul. 10 (1974) 77–86.
19] D. Pitre, Iodamide, in: K. Florey (Ed.), Analytical Profiles of
Drug Substances, vol. 15, Academic Press, New York, 1986, p.
3] P.P.T. Sah, S.A. Peoples, Isonicotinoyl hydrazones as antituber-
cular agents and derivatives for identification of aldeydes and
ketones, J. Am. Pharm. Assoc. XLIII (1954) 513–524.
4] D. Chakravarty, A. Bose, S. Bose, Synthesis and antitubercular
activity of isonicotinoyl and cyanoacetyl hydrazones, J. Pharm.
Sci. 53 (1964) 1036–1039.
5] M.G. Vigorita, R. Ottana, R. Maccari, F. Monforte, G. Bisi-
gnano, F.C. Pizzimenti, Synthesis and in vitro antimicrobial and
antitumoral screening of novel lipophilic isoniazid analogues. VI,
Boll. Chim. Farmaceutico 137 (1998) 267–276.
3
42.
20] K. Florey, Aztreonam, in: K. Florey (Ed.), Analytical Profiles of
Drug Substances, vol. 17, Academic Press, New York, 1988, p.
1
1.
21] A.M. Al-Obaid, F.J. Al-Shammary, K.A.M. Al-Rashood, M.S.
Mian, Furosemide, in: K. Florey (Ed.), Analytical Profiles of
Drug Substances, vol. 18, Academic Press, New York, 1989, p.
[
6] J.M. Patel, M.P. Dave, N.A. Langalia, K.A. Thaker, Studies
antitubercular and antibacterial agents: preparation of 1-(4-
aminobenzoyl)-2-benzalhydrazine and 1-[4-(phenylthioureido)-
benzoyl]-2-substituted benzalhydrazine, J. Indian Chem. Soc. 61
1
64.
[
22] M.A. Abounassif, N.A. Mian, M.S. Mian, Ceftazidime, in: K.
Florey (Ed.), Analytical Profiles of Drug Substances, vol. 19,
Academic Press, New York, 1990, p. 106.
(1984) 718–720 ref. C.A. 103 (1985) 87603r.
[
7] M.K. Jani, N.K. Undavia, P.B. Triverdi, Synthesis of 3-(p-dia-
zobenzoyl-N-b-arylidinohydrazone)indoles, J. Indian Chem. Soc.
[
23] S¸ .G. K o¨ m u¨ rc u¨ , S. Rollas, M. Ulgen, J.W. Gorrod, A. C¸ evikbas,
8
Evaluation of some arylhydrazones of p-amimobenzoic acid
hydrazide as antimicrobial agents and their in vitro hepatic
microsomal metabolism, Boll. Chim. Farmaceutico 134 (1995)
6
7 (1990) 601–602.
[
8] A.E.W. Smith, Preparation of some new 4-substituted deriva-
tives of p-amino-o-hydroxyphenyl-1,3,4-oxadiazolthiones-5 and
study of their mycobacteriostatic activity, Arz. Forsch. 17 (1967)
3
75–379.
[
[
24] S. Rollas, Synthesis of 3H-1,3,4-oxadiazoline-2-thione deriva-
tives and reductive cleavage with hydrazine hydrate, J. Fac.
Pharm. Istanbul. 19 (1983) 83–88.
1
150–1151.
[
9] A.C. Glasser, R.M. Doughty, Substituted heterocyclic thioureas
I. Antitubercular activity, J. Pharm. Sci. 51 (1962) 1031–1033.
25] S. Rollas, N. Kalyoncuo g˘ lu, D. S u¨ r-Altiner, Y. Ye g˘ eno g˘ lu,
[
[
10] L. Bukowski, M. Janowiec, 1-Methyl-1H-2-imidazo[4,5-
b]pyridinecarboxylic acid and some of its derivatives with sus-
pected antituberculotic activity, Pharmazie 51 (1996) 27–30.
5
-(4-Aminophenyl)
×-4-substituted-2,4-dihydro-3H-1,2,4-triazole-3-thiones, Phar-
mazie 48 (1993) 308–309.
11] I
:
. K u¨ c¸ u¨ kg u¨ zel, S¸ .G. K u¨ c¸ u¨ kg u¨ zel, S. Rollas, M. Kiraz, Some
-thioxo/alkylthio-1,2,4-triazoles with a substituted thiourea as
possible antimycobacterials, Biorg. Med. Chem. Letts. 11 (2001)
703–1707.
12] S¸ .G. K u¨ c¸ u¨ kg u¨ zel, S. Rollas, I
and antimycobacterial activity of some coupling products from
3
[26] S¸ .G. K u¨ c¸ u¨ kg u¨ zel, S. Rollas, H. Erdeniz, M. Kiraz, Synthesis,
characterization and antimicrobial evaluation of ethyl 2-arylhy-
drazono-3-oxobutyrates, Eur. J. Med. Chem. 34 (1999) 153–160.
[27] N. Ergen c¸ , S. Rollas, Some azopyrazoles, J. Fac. Pharm. Istan-
bul. 11 (1975) 138–157.
[28] U. Wrzeciono, K. Piethiewicz, B. Krzysztofik, W.D. Michalska,
M. Drozdowski, Sulfonamide. 11. Mitteilung: sulfonamid-
derivate des 1-methylphenyl-3-methylpyrazolons-(5) und 1-
methylphenyl-3,5-dimethylpyrazols, Pharmazie 33 (1978)
266–268.
[29] L. Collins, S.G. Franzblau, Microplate Alamar Blue Assay
versus BACTEC 460 system for high-thoughput screening of
compounds against Mycobacterium tuberculosis and Mycobac-
terium a6ium, Antimicrob. Agents Chemother. 41 (1997) 1004–
1009.
1
[
[
:
. K u¨ c¸ u¨ kg u¨ zel, M. Kiraz, Synthesis
4
1
-aminobenzoic acid hydrazones, Eur. J. Med. Chem. 34 (1999)
093–1099.
13] S¸ .G. K u¨ c¸ u¨ kg u¨ zel, S. Rollas, H. Erdeniz, M. Kiraz, A.C. Ekinci,
A. Vidin, Synthesis, characterization and pharmacological prop-
erties of some 4-arylhydazono-2-pyrazolne-5-one derivatives ob-
tained from heterocyclic amines, Eur. J. Med. Chem. 35 (2000)
7
61–771.
[
14] W.U. Malik, H.G. Garg, V. Arora, Synthesis and biological
evaluation of some 4-arylazopyrazoles, J. Pharm. Sci. 60 (1971)
1
738–1740.