Synthesis and antimycobacterial activities of some new N-acylhydrazone and thiosemicarbazide derivatives of 6-methyl-4,5-dihydropyridazin-3(2H)-one

Article abstract of DOI:10.1007/s00044-011-9770-6

In this study, twenty-five new 6-methyl-4, 5-dihydropyridazin-3(2H)-one derivatives having N-acylhydrazone and thiosemicarbazide moieties were synthesized. The target compounds were tested for their antimycobacterial activity in vitro against Mycobacteriu

Full text of DOI:10.1007/s00044-011-9770-6

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2012) 21:2388–2394
DOI 10.1007/s00044-011-9770-6
ORIGINAL RESEARCH
Synthesis and antimycobacterial activities of some new
N-acylhydrazone and thiosemicarbazide derivatives
of 6-methyl-4,5-dihydropyridazin-3(2H)-one
•
Oya Unsal Tan Keriman Ozadali
•
•
•
Perumal Yogeeswari Dharmarajan Sriram
Ayla Balkan
Received: 11 March 2011 / Accepted: 27 July 2011 / Published online: 7 August 2011
Ó Springer Science+Business Media, LLC 2011
Abstract In this study, twenty-five new 6-methyl-4,
5-dihydropyridazin-3(2H)-one derivatives having N-acy-
lhydrazone and thiosemicarbazide moieties were synthe-
sized. The target compounds were tested for their
antimycobacterial activity in vitro against Mycobacterium
tuberculosis H37Rv using the agar dilution method. Among
the synthesized compounds, N0-(2,4-dichlorobenzylidene)-
2-(3-methyl-6-oxo-5,6-dihydropyridazin-1(4H)-yl)acetohyd-
razide 4g was found to be the most active compound with
minimum inhibitory concentration of 0.78 lM and was
more potent than ethambutol and ciprofloxacin.
rifampin, isoniazid (INH), and ethambutol) and toxicities
of these compounds have further complicated the problem,
which clearly indicates the need for more effective, faster
acting chemotherapeutics with the lower toxicity for the
efficient management of tuberculosis.
Within the scope of antitubercular drug design, a num-
ber of N-acylhydrazones (NAH) having different hetero-
cyclic systems, such as thiadiazole, triazole, quinoline,
pyrazine, and imidazo[4,5-d]pyridine have been reported
as active compounds (Abdel-Aziz and Abdel-Rahman,
2010; Bukowski and Janowiec, 1996; Mamolo, et al., 2001;
Nayyar et al., 2007; Ulusoy et al., 2001). In the light of
these studies, it can be assumed that NAH moiety is a
suitable pharmacophore group for the treatment of tuber-
culosis. Thiosemicarbazides are less studied compared with
NAHs, however, there is an interesting report which claims
that some thiosemicarbazide derivatives of isoniazid show
higher activity than isoniazid (Cardia et al., 2006). On the
other hand, some compounds bearing pyridazine ring have
been reported to possess antimycobacterial activity (Islam
et al., 2008; Mantu et al. 2010).
Keywords Antimycobacterial activity ꢀ
6-Methyl-4,5-dihydropyridazin-3(2H)-one ꢀ
N-Acylhydrazone ꢀ Thiosemicarbazide
Introduction
Tuberculosis is an infection caused by Mycobacterium
tuberculosis and is the leading cause of infectious disease
mortality in the world (Dye, 2006). No new drugs have
been developed specifically against mycobacteria since the
1960s (Janin, 2007). The increase in M. tuberculosis strains
resistant to first-line antimycobacterial drugs (such as
On the basis of these findings, this study aimed to
investigate the synthesis and antimycobacterial activities
of some new 6-methyl-4,5-dihydropyridazin-3(2H)-one
derivatives having NAH and thiosemicarbazide moieties at
position 2 of the ring.
O. Unsal Tan (&) ꢀ K. Ozadali ꢀ A. Balkan
Department of Pharmaceutical Chemistry, Faculty of Pharmacy,
University of Hacettepe, Ankara, Turkey
Materials and methods
e-mail: oyaunsal@hacettepe.edu.tr
Chemistry
P. Yogeeswari ꢀ D. Sriram
Medicinal Chemistry & Antimycobacterial Research Laboratory,
Pharmacy Group, Birla Institute of Technology & Science –
Pilani, Hyderabad Campus, Jawahar Nagar,
Hyderabad 500 078, Andhra Pradesh, India
Melting points were determined using a Thomas-Hoover
Capillary Melting Point Apparatus (Philadelphia, PA,
USA) and are uncorrected. ATR-FTIR spectra were
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N0-Benzylidene-2-(3-methyl-6-oxo-5,6-dihydropyridazin-
1(4H)-yl)acetohydrazide (4a) Yield 51%, mp 197°C. IR;
1698 (C=O, pyridazinone), 1630 (C=O, hydrazide). 1H-
NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s; –CH3),
2.40–2.43 (2H; m; –CH2–), 2.49–2.51 (2H; m; –CH2–),
4.32 and 4.71 (2H; s; –CH2–CONH–), 7.41–7.69 (5H; m;
–C6H5), 7.98 and 8.21 (1H; s; –N=CH–), 11.46 and 11.54
(1H; s; –CO–NH–N=). ESI-MS (m/z); 295 [M?Na]?, 273
[M?H]? (100%).
obtained using the MIRacle ATR accessory (Pike tech-
nologies) in conjunction with a Spectrum BX FTIR spec-
trometer (Perkin Elmer) and were reported in cm-1. The
1H NMR spectra (DMSO-d6) were measured on a Varian
Mercury 400 FT NMR spectrophotometer using TMS as an
internal reference. Chemical shifts are reported in d values
(ppm). The ESI-MS spectra were recorded on a micromass
ZQ-4000 single quadruple mass spectrometer. Elemental
analyses (C, H, N) were performed on Leco CHNS 932
analyzer. The results were within 0.4% of the theoretical
values.
0
N -(4-Chlorobenzylidene)-2-(3-methyl-6-oxo-5,6-dihydro-
pyridazin-1(4H)-yl)acetohydrazide (4b) Yield 55%, mp
190°C. IR; 1683 (C=O, pyridazinone), 1656 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.94 (3H; s;
–CH3), 2.35–2.40 (2H; m; –CH2–), 2.46–2.50 (2H; m;
–CH2–), 4.29 and 4.68 (2H; s; –CH2–CONH–), 7.45–7.50
(2H; m; –C6H4–Cl, H3, H5), 7.67–7.70 (2H; m; –C6H4–Cl,
H2, H6), 7.94 and 8.18 (1H; s; –N=CH–), 11.51 and 11.57
(1H; s; –CO–NH–N=). ESI-MS (m/z); 331 [M?Na?2]?,
329 [M?Na]?, 307 [M?H]? (100%).
6-Methyl-4,5-dihydropyridazin-3(2H)-one (1)
This compound was prepared according to the method
described in the literature (Reichelt and Reissig, 1984).
Ethyl 2-(3-methyl-6-oxo-5,6-dihydropyridazin-1(4H)-
yl)acetate (2)
The mixture of 1(0.05 mol) and anhydrous potassium
carbonate (0.075 mol) in acetone (100 ml) was refluxed for
4 h. To the mixture, a solution of 0.05 mol ethyl bromo-
acetate in acetone (25 ml) was added dropwise. After
refluxing for 2 h, the reaction mixture was poured into ice-
cold water (250 ml) and extracted with ethyl acetate. The
oily residue obtained was used in the next step without
further purification.
N0-(4-Bromobenzylidene)-2-(3-methyl-6-oxo-5,6-dihydro-
pyridazin-1(4H)-yl)acetohydrazide (4c) Yield 80%, mp
211°C. IR; 1688 (C=O, pyridazinone), 1656 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s;
–CH3), 2.40–2.42 (2H; m; –CH2–), 2.49–2.51 (2H; m;
–CH2–), 4.32 and 4.70 (2H; s; –CH2–CONH–), 7.61–7.66
(4H; m; –C6H4–Br), 7.95 and 8.18 (1H; s; –N=CH–), 11.54
and 11.60 (1H; s; –CO–NH–N=). ESI–MS (m/z); 375
[M?Na?2]?, 373 [M?Na]?, 351 [M?H]? (100%).
2-(3-Methyl-6-oxo-5,6-dihydropyridazin-1-(4H)-
yl)acetohydrazide (3)
N0-(4-Methylbenzylidene)-2-(3-methyl-6-oxo-5,6-dihydropyri-
dazin-1(4H)-yl)acetohydrazide (4d) Yield 55%, mp
193°C. IR; 1666 (C=O, pyridazinone, hydrazide). 1H-NMR
(DMSO-d6, 400 MHz); d 1.97 (3H; s; –CH3), 2.33 (3H; s;
ar-CH3), 2.40–2.42 (2H; m; –CH2–), 2.48–2.51 (2H; m;
–CH2–), 4.31 and 4.69 (2H; s; –CH2–CONH–), 7.23–7.26
(2H; m; ar, H3, H5), 7.56–7.59 (2H; m; ar, H2, H6), 7.94
and 8.16 (1H; s; –N=CH–), 11.39 and 11.46 (1H; s; –CO–
NH–N=). ESI-MS (m/z); 309 [M?Na]?, 287 [M?H]?
(100%).
Compound 2 (0.01 mol) was refluxed with hydrazine
(0.20 mol) in 50 ml of absolute ethanol for 1 h. The sol-
vent was evaporated under reduced pressure and the resi-
due was crystallized from ethanol to give 3. mp 194°C. IR;
3306, 3210 (N–H), 1667 (C=O, ester), 1644 (C=O, pyrid-
azinone). 1H-NMR (DMSO-d6, 400 MHz); 1.95 (3H; s;
–CH3), 2.35–2.40 (2H; m; –CH2–), 2.49–2.53 (2H; m;
–CH2–), 4.15 (2H; s; –CH2–CONH–), 4.21 (2H; b;
–NHNH2), 9.02 (1H; b; –NHNH2). ESI-MS (m/z); 207
[M?Na]?, 185 [M?H] (100%).
N0-(4-Methoxybenzylidene)-2-(3-methyl-6-oxo-5,6-dihyd-
ropyridazin-1(4H)-yl)acetohydrazide (4e) Yield 54%, mp
185°C. IR; 1673 (C=O, pyridazinone), 1651 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s;
–CH3), 2.40–2.42 (2H; m; –CH2–), 2.48–2.51 (2H; m;
–CH2–), 3.79 (3H; s; ar-OCH3), 4.30 and 4.68 (2H; s;
–CH2–CONH–), 6.98–7.02 (2H; m; ar, H3, H5), 7.61–7.64
(2H; m; ar, H2, H6), 7.92 and 8.15 (1H; s; –N=CH–), 11.31
and 11.39 (1H; s; –CO–NH–N=). ESI-MS (m/z); 325
[M?Na]?, 203 [M?H]? (100%).
General procedure for the preparation
of N-acylhydrazones (4a–m and 5a–h)
To a stirred mixture of 0.01 mol hydrazide (3) in 20 ml of
ethanol, 0.01 mol of appropriate aldehyde or acetophenone
was added and stirring was continued for 2 h in the pres-
ence of HCl (2 drops). The mixture was poured into cold
water and neutralized with 10% aqueous sodium bicar-
bonate solution. The obtained solid was filtered, washed
with water, and crystallized from acetonitrile.
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N0-(4-Nitrobenzylidene)-2-(3-methyl-6-oxo-5,6-dihydro-
pyridazin-1(4H)-yl)acetohydrazide (4f) Yield 94%, mp
216°C. IR; 1687 (C=O, pyridazinone), 1640 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s;
–CH3), 2.41–2.43 (2H; m; –CH2–), 2.50–2.51 (2H; m;
–CH2–), 4.36 and 4.76 (2H; s; –CH2–CONH–), 7.95–8.32
(5H; m; ar, H2, H3, H5, H6, –N=CH–), 11.80 and 11.84
(1H; s; –CO–NH–N=). ESI-MS (m/z); 340 [M?Na]?, 318
[M?H]? (100%).
N0-(Furan-2-ylmethylene)-2-(3-methyl-6-oxo-5,6-dihyd-
ropyridazin-1(4H)-yl)acetohydrazide (4k) Yield 43%,
mp 199°C. IR; 1692 (C=O, pyridazinone), 1661 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s;
–CH3), 2.39–2.42 (2H; m; –CH2–), 2.48–2.51 (2H; m;
–CH2–), 4.30 and 4.62 (2H; s; –CH2–CONH–), 6.61 (1H;
d; furan, H4), 6.89 (1H; t; furan, H3), 7.82 (1H; d; furan,
H5), 7.86 and 8.09 (1H; s; –N=CH–), 11.39 and 11.48 (1H;
s; –CO–NH–N=). ESI-MS (m/z); 285 [M?Na]?, 263
[M?H]? (100%).
N0-(2,4-Dichlorobenzylidene)-2-(3-methyl-6-oxo-5,6-di-
hydropyridazin-1(4H)-yl)acetohydrazide
(4g) Yield
N0-(Thiophen-2-ylmethylene)-2-(3-methyl-6-oxo-5,6-dihyd-
ropyridazin-1(4H)-yl)acetohydrazide (4l) Yield 73%, mp
209°C. IR; 1694 (C=O, pyridazinone), 1626 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s;
–CH3), 2.39-2.42 (2H; m; –CH2–), 2.48–2.51 (2H; m;
–CH2–), 4.30 and 4.61 (2H; s; –CH2–CONH–), 7.10–7.14
(1H; m; thiophene, H4), 7.42–7.46 (1H; m; thiophene, H3),
7.63–7.66 (1H; m; thiophene, H5), 8.15 and 8.43 (1H; s;
–N=CH–), 11.41 and 11.52 (1H; s; –CO–NH–N=). ESI-MS
(m/z); 301 [M?Na]?, 279 [M?H]? (100%).
62%, mp 203°C. IR; 1694 (C=O, pyridazinone), 1640
(C=O, hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97
(3H; s; –CH3), 2.40–2.42 (2H; m; –CH2–), 2.49–2.52 (2H;
m; –CH2–), 4.33 and 4.73 (2H; s; –CH2–CONH–), 7.48
(1H; d; ar, H5, J: 8.8 Hz), 7.72 (1H; s; ar, H3), 7.99 (1H; d;
ar, H6, J: 8.8 Hz), 8.31 and 8.56 (1H; s; –N=CH–), 11.77
(1H; s; –CO–NH–N=). ESI-MS (m/z); 365 [M?Na?2]?,
363 [M?Na]?, 341 [M?H]? (100%).
N0-(4-(Trifluoromethyl)benzylidene)-2-(3-methyl-6-oxo-5,
6-dihydropyridazin-1(4H)-yl)acetohydrazide (4h) Yield
59%, mp 165°C. IR; 1688 (C=O, pyridazinone), 1639
(C=O, hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97
(3H; s; –CH3), 2.38–2.42 (2H; m; –CH2–), 2.48–2.50 (2H;
m; –CH2–), 4.37 and 4.74 (2H; s; –CH2–CONH–),
7.30–7.53 (2H; m; ar, H3, H5), 7.79–7.84 (2H; m; ar, H2,
H6), 8.05 and 8.28 (1H; s; –N=CH–), 11.73 (1H; s; –CO–
NH–N=). ESI-MS (m/z); 363 [M?Na]?, 341 [M?H]?
(100%).
N0-(4-Ethoxybenzylidene)-2-(3-methyl-6-oxo-5,6-dihydro-
pyridazin-1(4H)-yl)acetohydrazide (4m) Yield 64%, mp
166°C. IR; 1679 (C=O, pyridazinone), 1643 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.35 (3H; t;
–OCH2–CH3), 1.97 (3H; s; –CH3), 2.39–2.41 (2H; m;
–CH2–), 2.48–2.51 (2H; m; –CH2–), 4.08 (3H; q; –OCH2–
CH3), 4.30 and 4.68 (2H; s; –CH2–CONH–), 7.59–7.62
(2H; m; ar, H3, H5), 7.78–7.81 (2H; m; ar, H2, H6), 7.91
and 8.14 (1H; s; –N=CH–), 11.31 and 11.39 (1H; s; –CO–
NH–N=). ESI-MS (m/z); 339 [M?Na]?, 317 [M?H]?
(100%).
N0-(4-(Dimethylamino)benzylidene)-2-(3-methyl-6-oxo-5,6-di-
hydropyridazin-1(4H)-yl)acetohydrazide (4i) Yield 67%,
mp 200°C. IR; 1698 (C=O, pyridazinone), 1646 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.94 (3H; s;
–CH3), 2.37–2.39 (2H; m; –CH2–), 2.46–2.49 (2H; m;
–CH2–), 2.94 (6H; s; N(CH3)2), 4.25 and 4.63 (2H; s;
–CH2–CONH–), 6.69–6.73 (2H; m; ar, H3, H5), 7.44–7.48
(2H; m; ar, H2, H6), 7.81 and 8.02 (1H; s; –N=CH–), 11.10
and 11.20 (1H; s; –CO–NH–N=). ESI-MS (m/z); 338
[M?Na]?, 316 [M?H]? (100%).
N0-(1-Phenylethylidene)-2-(3-methyl-6-oxo-5,6-dihydropy-
ridazin-1(4H)-yl)acetohydrazide (5a) Yield 50%, mp
171°C. IR; 1685 (C=O, pyridazinone), 1661 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s;
–CH3), 2.25 and 2.27 (3H; s; –N=C–CH3), 2.38–2.42 (2H;
m; –CH2–), 2.49–2.51 (2H; m; –CH2–), 4.44 and 4.74 (2H;
s; –CH2–CONH–), 7.39–7.80 (5H; m; ar), 10.48 and 10.76
(1H; s; –CO–NH–N=). ESI-MS (m/z); 309 [M?Na]?, 287
[M?H]? (100%).
N0-(4-Fluorobenzylidene)-2-(3-methyl-6-oxo-5,6-dihydro-
pyridazin-1(4H)-yl)acetohydrazide (4j) Yield 58%, mp
180°C. IR; 1701 (C=O, pyridazinone), 1650 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.96 (3H; s;
–CH3), 2.38–2.43 (2H; m; –CH2–), 2.51–2.52 (2H; m;
–CH2–), 4.35 and 4.70 (2H; s; –CH2–CONH–), 7.25–7.29
(2H; m; ar, H3, H5), 7.73–7.78 (2H; m; ar, H2, H6), 7.99
and 8.25 (1H; s; –N=CH–), 11.18 and 11.57 (1H; s; –CO–
NH–N=). ESI–MS (m/z); 313 [M?Na]?, 291 [M?H]?
(100%).
N0-(1-(4-Chlorophenyl)ethylidene)-2-(3-methyl-6-oxo-5,6-
dihydropyridazin-1(4H)-yl)acetohydrazide (5b) Yield 66%,
mp 212°C. IR; 1660 (C=O, pyridazinone), 1643 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s;
–CH3), 2.24 and 2.26 (3H; s; –N=C–CH3), 2.38–2.42 (2H;
m; –CH2–), 2.49–2.51 (2H; m; –CH2–), 4.44 and 4.74 (2H;
s; –CH2–CONH–), 7.45–7.59 (2H; m; ar, H3, H5),
7.80–7.82 (2H; m; ar, H2, H6), 10.53 and 10.82 (1H; s; –
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Med Chem Res (2012) 21:2388–2394
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CO–NH–N=). ESI-MS (m/z); 345 [M?Na?2]?, 343
[M?Na]?, 321 [M?H]? (100%).
4.44 and 4.78 (2H; s; –CH2–CONH–), 7.40 (1H; d; ar, H5,
J: 8.8 Hz), 7.71 (1H; s; ar, H3), 7.96 (1H; d; ar, H6, J:
8.8 Hz), 10.73 and 10.96 (1H; s; –CO–NH–N=). ESI-MS
(m/z); 379 [M?Na?2]?, 377 [M?Na]?, 355 [M?H]?
(100%).
N0-(1-(4-Bromophenyl)ethylidene)-2-(3-methyl-6-oxo-5,6-
dihydropyridazin-1(4H)-yl)acetohydrazide (5c) Yield 63%,
1
mp 225°C. IR; 1660 (C=O, pyridazinone, hydrazide). H-
NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s; –CH3), 2.23
and 2.26 (3H; s; –N=C–CH3), 2.38–2.42 (2H; m; –CH2–),
2.49–2.51 (2H; m; –CH2–), 4.44 and 4.73 (2H; s; –CH2–
CONH–), 7.58–7.63 (2H; m; ar, H3, H5), 7.73–7.75 (2H; m;
ar, H2, H6), 10.53 and 10.83 (1H; s; –CO–NH–N=). ESI-
MS (m/z); 389 [M?Na?2]?, 387 [M?Na]?, 365 [M?H]?
(100%).
N0-(1-(4-Phenylphenyl)ethylidene)-2-(3-methyl-6-oxo-5,6-
dihydropyridazin-1(4H)-yl)acetohydrazide (5h) Yield 70%,
mp 221°C. IR; 1683 (C=O, pyridazinone), 1661 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.98 (3H; s;
–CH3), 2.28 and 2.31(3H; s; –N=C–CH3), 2.40–2.43 (2H; m;
–CH2–), 2.50–2.54 (2H; m; –CH2–), 4.45 and 4.77 (2H; s;
–CH2–CONH–), 7.37–7.89 (9H; m; ar), 10.51 and 10.81
(1H; s; –CO–NH–N=). ESI-MS (m/z); 385 [M?Na]?, 363
[M?H]? (100%).
N0-(1-(4-Methylphenyl)ethylidene)-2-(3-methyl-6-oxo-5,6-di-
hydropyridazin-1(4H)-yl)acetohydrazide (5d) Yield 67%,
mp 211°C. IR; 1683 (C=O, pyridazinone), 1658 (C=O,
hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97 (3H; s;
–CH3), 2.22 and 2.24 (3H; s; –N=C–CH3), 2.32 (3H; s; ar-
CH3), 2.38–2.42 (2H; m; –CH2–), 2.49–2.51 (2H; m;
–CH2–), 4.42 and 4.72 (2H; s; –CH2–CONH–), 7.20–7.22
(2H; d; ar, H3, H5, J:8.4 Hz), 7.67–7.69 (2H; d; ar, H2, H6,
J:8.4 Hz), 10.42 and 10.70 (1H; s; –CO–NH–N=). ESI-MS
(m/z); 323 [M?Na]?, 301 [M?H]? (100%).
General procedure for the preparation
of thiosemicarbazides (6a–c)
An equimolar amount of appropriate isothiocyanate was
added to a solution of hydrazide 6a–c in 50 ml of ethanol.
The reaction mixture was refluxed for 4 h and cooled to
room temperature. The precipitated solid was filtered,
washed with water, and purified by crystallization from
ethyl acetate.
N0-(1-(4-Methoxyphenyl)ethylidene)-2-(3-methyl-6-oxo-5,
6-dihydropyridazin-1(4H)-yl)acetohydrazide (5e) Yield
56%, mp 185°C. IR; 1675 (C=O, pyridazinone), 1659
(C=O, hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.97
(3H; s; –CH3), 2.21 and 2.23 (3H; s; –N=C–CH3),
2.38–2.42 (2H; m; –CH2–), 2.49–2.50 (2H; m; –CH2–),
3.79 (3H; s; ar-OCH3), 4.41 and 4.72 (2H; s; –CH2–
CONH–), 6.94–6.98 (2H; m; ar, H3, H5), 7.72–7.75 (2H; m;
ar, H2, H6), 10.38 and 10.65 (1H; s; –CO–NH–N=). ESI-
MS (m/z); 339 [M?Na]?, 317 [M?H]? (100%).
4-Methyl-1-[2-(3-methyl-6-oxo-5,6-dihydropyridazin-1(4H)-
yl)acetyl]thiosemicarbazide (6a) Yield 52%, mp 198°C.
IR; 1724 (C=O, pyridazinone), 1634 (C=O, hydrazide),
1
1201 (C=S). H-NMR (DMSO-d6, 400 MHz); d 1.96 (3H;
s; –CH3), 2.39–2.41 (2H; m; –CH2–), 2.47–2.51 (2H; m;
–CH2–), 2.86 (3H; d; NH–CH3), 4.31 (2H; s; –CH2–
CONH–), 7.81 (1H; br; NH–CH3), 9.30 (1H; br; –NH–NH–
CS), 9.86 (1H; br; –NH–NH–CS). ESI-MS (m/z); 280
[M?Na]?, 258 [M?H]? (100%).
N0-(1-(4-Nitrophenyl)ethylidene)-2-(3-methyl-6-oxo-5,6-di-
hydropyridazin-1(4H)-yl)acetohydrazide (5f) Yield 88%,
mp[270°C. IR; 1672 (C=O, pyridazinone, hydrazide). 1H-
NMR (DMSO-d6, 400 MHz); d 1.98 (3H; s; –CH3), 2.31
and 2.42 (3H; s; –N=C–CH3), 2.40–2.44 (2H; m; –CH2–),
2.50–2.52 (2H; m; –CH2–), 4.47 and 4.78 (2H; s; –CH2–
CONH–), 8.03–8.07 (2H; m; ar, H2, H6,), 8.23–8.29 (2H;
m; ar, H3, H5), 10.72 and 10.98 (1H; s; –CO–NH–N=).
ESI-MS (m/z); 354 [M?Na]?, 332 [M?H]? (100%).
4-Ethyl-1-[2-(3-methyl-6-oxo-5,6-dihydropyridazin-1(4H)-
yl)acetyl]thiosemicarbazide (6b) Yield 65%, mp 190°C.
IR; 1727 (C=O, pyridazinone), 1634 (C=O, hydrazide),
1
1192 (C=S). H-NMR (DMSO-d6, 400 MHz); d 1.06 (3H;
t; –NH–CH2–CH3), 1.96 (3H; s; –CH3), 2.37–2.41 (2H; m;
–CH2–), 2.48–2.51 (2H; m; –CH2–), 3.43–3.46 (2H; m;
–NH–CH2–CH3), 4.29 (2H; s; –CH2–CONH–), 7.73 (1H;
br; NH–CH2–CH3), 9.26 (1H; br; –NH–NH–CS), 9.87 (1H;
br; –NH–NH–CS). ESI-MS (m/z); 294 [M?Na]?, 272
[M?H]? (100%).
N0-(1-(2,4-Dichlorophenyl)ethylidene)-2-(3-methyl-6-oxo-5,
6-dihydropyridazin-1(4H)-yl)acetohydrazide (5g) Yield
72%, mp 192°C. IR; 1682 (C=O, pyridazinone), 1660
(C=O, hydrazide). 1H-NMR (DMSO-d6, 400 MHz); d 1.98
(3H; s; –CH3), 2.33 and 2.39 (3H; s; –N=C–CH3),
2.41–2.43 (2H; m; –CH2–), 2.50–2.53 (2H; m; –CH2–),
4-Phenyl-1-[2-(3-methyl-6-oxo-5,6-dihydropyridazin-1(4H)-yl)
acetyl]thiosemicarbazide (6c) Yield 74%, mp 158°C. IR;
1703 (C=O, pyridazinone), 1645 (C=O, hydrazide), 1200
(C=S). 1H-NMR (DMSO-d6, 400 MHz); d 1.96 (3H; s;
–CH3), 2.41–2.42 (2H; m; –CH2–), 2.49–2.50 (2H; m;
123

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Med Chem Res (2012) 21:2388–2394
–CH2–), 4.36 (2H; s; –CH2–CONH–), 7.15–7.47 (5H; m;
NH–C6H5), 9.41 (1H; br; NH–C6H5), 9.71 (1H; br; –NH–
NH–CS), 10.14 (1H; br; –NH–NH–CS). ESI–MS (m/z);
342 [M?Na]?, 320 [M?H]? (100%).
treatment of 1 with ethyl bromoacetate in the presence of
potassium carbonate. The reaction of 2 with excess of
hydrazine hydrate gave 2-(3-Methyl-6-oxo-5,6-dihydropy-
ridazin-1-(4H)-yl)acetohydrazide 3. Compound 3 was
treated with the appropriate benzaldehydes and acetophe-
nones in the presence of HCl to synthesize 4a–m and 5a–h,
respectively. Finally, thiosemicarbazide derivatives 6a–c
were prepared by refluxing 3 with the appropriate
isothiocyanates.
Evaluation of antimycobacterial activity
The target compounds were screened for their in vitro
antimycobacterial activity against M. tuberculosis, in
Middlebrook 7H11 agar medium supplemented with
OADC by agar dilution method similar to that recom-
mended by the National Committee for Clinical Laboratory
Standards for the determination of MIC in triplicate (1995).
The minimum inhibitory concentration (MIC) was defined
as the minimum concentration of the compound required to
provide complete inhibition of bacterial growth.
The structures of the target compounds 3 and 4a–6c
1
were characterized using spectral methods (IR, H-NMR,
and ESI-MS). The bands at 3306 and 3210 cm-1 in the IR
spectra and the signals at 4.21 and 9.02 ppm in the
1H-NMR spectra of 3 were the evidences of the presence
of hydrazide structure. N-Acylhydrazones may exist as
geometrical isomers (E/Z) with respect to the C=N dou-
ble bonds and as rotamers (cis/trans) about amide
N–C(O) bond (Palla et al., 1986; Syakaev et al., 2006). As
mentioned in our previous article (Unsal-Tan et al., 2010),
in the 1H-NMR spectra of 4a–m and 5a–h, the double
signals were observed due to the presence of cis and trans
isomers. In the 1H-NMR spectra of 4a–m, the signals
belonging to methylidene and imine protons were observed
at around 4.30 (trans), 4.70 (cis) and 8.20 (trans), 7.90
Results and discussion
Chemistry
The target compounds (3, 4a–6c) were synthesized via the
route outlined in Scheme 1. The starting compound,
6-methyl-4,5-dihydropyridazin-3(2H)-one 1 was prepared
by the reaction of levulinic acid with hydrazine hydrate
according to the method reported earlier (Reichelt and
Reissig, 1984). Although 2 and 3 are already known
(Scifinder), the synthetic methods and spectral data of them
are also given in this study. Ethyl 2-(3-methyl-6-oxo-5,6-
dihydropyridazin-1(4H)-yl)acetate 2 was prepared by the
1
(cis) ppm, respectively. Different from 4a–m, in the H-
NMR spectra of 5a–h, the methyl protons were observed as
double signals at around 2.20 (trans) and 2.30 (cis) ppm in
place of imine proton. The signals of the amide protons in
4a–m and 5a–h were also seen at around 11–12 ppm as
double singlets with some exceptions. The strong absorp-
tion band at around 1200 cm-1 due to C=S stretching in the
IR and three broad signals at around 7.70, 9.30, and
1
O
O
9.90 ppm in the H-NMR spectra of 6a–c exhibited the
presence of thiosemicarbazide structure. Furthermore, the
structures of all the target compounds were confirmed by
the peaks belonging to [M?Na]? and [M?H]? seen in the
ESI mass spectra.
ii
i
NH
N
NCH2COOC2H5
N
CH3COCH2CH2COOH
CH3
1
CH3
2
O
O
Antimycobacterial activity
R1
C
iv
iii
NCH2CONHN
N
R2
NCH2CONHNH2
N
The synthesized compounds 3 and 4a–6c were evaluated
for in vitro antimycobacterial activity against M. tubercu-
losis by agar dilution method using Middlebrook 7H11
agar medium supplemented with OADC. The results for
the Minimum Inhibitory Concentrations (MICs) are pre-
sented in Table 1.
CH3
CH3
4a-m, 5a-h
O
3
v
S
NCH2CONHNH
C
NH R3
N
When the results were examined, benzaldehyde-derived
NAH compounds showed higher antimycobacterial activity
than their acetophenone-derived counterparts with some
exceptions. The semicarbazide derivatives 6a–c were
found as the least active compounds in the series.
In general, the benzaldehyde-derived NAH compounds
showed remarkable antimycobacterial activity except 4k,
CH3
6a-c
Scheme 1 Synthesis of the compounds. Reagents and conditions:
(i) NH2NH2.H2O, reflux; (ii) BrCH2COOC2H5, K2CO3, reflux; (iii)
NH2NH2.H2O, reflux; (iv) appropriate aldehydes and acetophenones,
rt; (v) R3NCS, reflux
123

Med Chem Res (2012) 21:2388–2394
2393
and 4 h with MIC of 6.25 and 1.56 lM, respectively) on the
phenyl ring increased the antimycobacterial activity while
electron donating groups (CH3, OCH3 N(CH3)2, and OC2H5)
diminished the activity (4d, 4e, 4i, and 4m with MIC
C12.5 lM).
It is noteworthy that the acetophenone-derived NAH
compounds 5a–h were devoid of antimycobacterial activity
(C12.5 lM). This observation may be due to the intro-
duction of a bulky group (methyl) in the C=N double bond
of NAH moiety. Among the 5a–h, compounds substituted
with Cl, Br, NO2 (5b, 5c, 5f, and 5g with MIC of 12.5 lM)
are more active than rest of the series.
Table 1 Physical data and antimycobacterial activity of synthesized
compounds
Compound
R1
R2
R3
Yield M.p. MIC
(lM)
(%)
(°C)
3
–
–
–
51
55
80
55
54
94
62
59
67
58
43
73
64
50
66
63
67
56
88
72
70
194 25
4a
H
H
H
H
H
H
H
H
H
H
H
H
H
CH3
H
197 12.5
190 3.13
211 1.56
193 12.5
185 25
216 6.25
203 0.78
165 1.56
200 12.5
180 1.56
199 [25
209 25
166 [25
171 25
212 12.5
225 12.5
211 25
185 25
[270 12.5
192 12.5
221 25
198 [25
190 [25
158 [25
0.05
4b
Cl
4c
Br
4d
CH3
OCH3
NO2
2,4-Cl
CF3
4e
4f
4g
4h
Acknowledgments The authors thank the financial support pro-
vided by Scientific Research Fund of Hacettepe University through
Project 0801301003. The authors also thank Assoc. Prof Dr. Aymil
Dogan (Faculty of Letters, Translation and Interpretation Department,
Hacettepe University) for grammatical revision.
4i
N(Me)2
F
4j
4k
Furfural
Thiophene
OC2H5
H
4l
Conflict of interests The authors report no declarations of interest.
4m
5a
5b
CH3 Cl
5c
CH3 Br
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