Synthesis and antibacterial activity of 4-benzoyl-1-methyl-5-phenyl-1H- pyrazole-3-carboxylic acid and derivatives

Article abstract of DOI:10.1016/j.farmac.2004.09.003

Some new 1H-pyrazole-3-carboxylic acid and pyridazinone derivatives were synthesized and evaluated for their antibacterial activities against Bacillus cereus ATCC 7064, Staphylococcus aureus ATCC 6538, Escherichia coli ATCC 4230 and Pseudomonas putida usi

Full text of DOI:10.1016/j.farmac.2004.09.003

IL FARMACO 60 (2005) 23–26
http://france.elsevier.com/direct/FARMAC/
Original article
Synthesis and antibacterial activity
of 4-benzoyl-1-methyl-5-phenyl-1H-pyrazole-3-carboxylic acid
and derivatives
a,
b
a
a
Esvet Akbas *, Ismet Berber , Ahmet Sener , Beybala Hasanov
a
Faculty of Arts and Sciences, Chemistry Department, Organic Chemistry Division, Yuzuncu Yıl University, 65080-Van-Turkey
Faculty of Arts and Sciences, Biology Department, Molecular Biology Division, Yuzuncu Yıl University, 65080-Van-Turkey
b
Received 1 February 2004; accepted 25 September 2004
Available online 19 December 2004
Abstract
Some new 1H-pyrazole-3-carboxylic acid and pyridazinone derivatives were synthesized and evaluated for their antibacterial activities
against Bacillus cereus ATCC 7064, Staphylococcus aureus ATCC 6538, Escherichia coli ATCC 4230 and Pseudomonas putida using tube
dilution method. The minimal inhibitory concentrations (MICs) experiments revealed that all chemical compounds showed inhibitor effects
on the growth of the test microorganisms. Moreover, the results of this research showed that the compound named as 5c was the best com-
pound in the series, exhibiting antibacterial activity against both Gram-positive and Gram-negative bacteria.
©
2004 Published by Elsevier SAS.
Keywords: Pyrazole; Pyridazine; Antibacterial activity
1
. Introduction
cal behavior of furandione 1 toward a aliphatichydrazine com-
pound, and present the reactions of cyclic oxalyl compounds
with methylhydrazine to prepare new pyrazole and pyridazine
derivatives, and theirs antibacterial activity.
In recent years, the number of life-threatening infectious
caused by multi-drug resistant Gram-positive and Gram-
negative pathogen bacteria have reached an alarming level in
many countries around the world [1,2]. A number of clinical
reports in the United States and worldwide have indepen-
dently described the emergence of vancomycin resistance in
methicillin-resistance Staphylococcus aureus (MRSA) iso-
lates and other human pathogen Gram-negative isolates [3].
Infections caused by these microorganisms pose a serious
challenge to the medical community and need for an effec-
tive therapy has led to a search for novel antibacterial agents.
Literatures report that pyrazole compounds, among their
numerous pharmacological properties, possess also antimi-
crobial activity [4,5]. Cyclic oxalyl compounds of type 1 have
been used successfully as starting materials to synthesize vari-
ous 1,4,5-trisubstitue pyrazol-3-carboxylic acids and their
derivatives via reaction with various hydrazines or hydra-
zones for about two decades [6–8]. Here, we report the chemi-
2. Chemistry
The furandione 1 was reacted with methylhydrazine to give
the 1H-pyrazole-3-carboxylic acid 2 together with a new
pyridazinone derivative 3, 2 was converted into the corre-
sponding amide 5a, ester 5b or urea 5c derivatives, respec-
tively, via reactions of its acid chloride 4 with alcohol or vari-
ous N-nucleophiles. Nitrile derivative 6 was obtained by
dehydration of 5a in a mixture of SOCl2 and DMF
(Scheme 1).
3
. Experimental
Solvents were dried by refluxing with the appropriate dry-
ing agents (metallic sodium for ether; CaCl , or Na SO for
*
Corresponding author.
2
2
4
E-mail address: esvakbas@hotmail.com (E. Akbas).
benzene, toluene...) and distilled before use. Melting points
0014-827X/$ - see front matter © 2004 Published by Elsevier SAS.
doi:10.1016/j.farmac.2004.09.003

24
E. Akbas et al. / IL FARMACO 60 (2005) 23–26
1
21.99, 42.56 ppm (N–CH ). Anal. calcd. for C H N O :
3 18 14 2 3
C, 70.58; H, 4.61; N, 9.15. Found: C, 70.57; H, 4.62; N, 9.16.
3
.3. 4-Benzoyl-1-methyl-5-phenyl-1H-pyrazole-3-carbonyl
chloride 4
Compound 2 (0.306 g, 1 mmole) and thionylchloride (1 ml,
3.8 mmoles) were refluxed on a steam bath for 5 h. After
1
cooling, the crude precipitate was isolated by filtration and
recrystallized from carbon tetrachloride, yield 0.216 g (68%),
–
1
mp 160 °C; IR: 1750 (C=O, acyl), 1669 cm (C=O, ben-
1
zoyl); H-NMR (CDCl ): d = 7.76–7.31 (m, 10H, Harom), 3.96
3
1
3
Scheme 1
.
Synthesis of compounds 2–6. Reagents: (a) CH NHNH ; (b)
(s, 3H, CH3); C-NMR (CDCl ): d = 192.13 (C=O, ben-
3
2
3
SOCl ; (c) NH ; (d) n-C -H OH; (e) Ph-NHCONH ; (f) DMF-SOCl .
2
3
4
9
2
2
zoyl), 163.27 (C=O, acyl), 147.69, (C ), 144.67 (C ), 139.25,
3
5
1
35.62, 132.11, 131.55, 131.31, 130.97, 130.54, 128.85
were determined on an Electrothermal Gallenkamp appara-
tus and are uncorrected. Microanalyses were performed on a
Carlo Erba Elemental Analyzer Model 1108. The IR spectra
were obtained in as potassium bromide pellets using a Matt-
(
C-Ph), 125.01 (C ), 40.73 (N–CH ). Anal. calcd. for
4
3
C H N O Cl: C, 66.55; H, 4.03; N, 8.63. Found: C, 65.50;
H, 4.05; N, 8.66.
1
8 13 2 2
1
13
son 1000 FTIR spectrometer. The H and C-NMR spectra
were recorded on Varian XL-200 (200 MHz) and Varian
XL-200 (50 MHz) spectrometers, respectively, using TMS
as an internal standard. All experiments were followed by
TLC using DC Alufolien Kieselgel 60 F 254 Merck and
Camag TLC lamp (254/366 nm).
3
.4. 4-Benzoyl-1-methyl-5-phenyl-1H-pyrazole-3-
carboxamide 5a
A moderate stream of gaseous ammonia was allowed to
bubble through a solution of pyrazole-3-carboxylic acid chlo-
ride 4 (0.324 g, 1 mmole) in 10 ml of carbon tetrachloride
during 30 min with ice-cooling. Then the crude precipitate
was filtered off and recystallized from methanol to give
3.1. 4-Benzoyl-1-methyl-5-phenyl-1H-pyrazole-3-carboxylic
acid 2
–1
0
.184 g (65%) of 5a, mp 220 °C; IR: 3451–3320 cm (NH),
–
1
–1
1
An equimolar mixture of furandione 1 (0.278 g, 1 mmole)
1662 cm (C=O, benzoyl), 1594 cm (C=O, amide). H-
and methylhydrazine (3.1 ml, 1 mmole) was mixture in room
temperature in dry benzene (30 ml) for approximately 60 min.
After the precipitate was filtered off and treated with dry ether
to give a crude solid that was recrystallized from methyl alco-
NMR(CDCl ): d 7.69–7.20 (m, 10H, Harom), 5.57–5.55 (bs,
3
13
2H, NH ), 3.89 (s, 3H, N–CH ). C-NMR (CDCl ): d 194.35
2
3
3
(C=O, benzoyl), 164.48 (C=O, amide), 147.52 (C ), 147.48
3
(C ), 145.85, 140.19, 134.82, 132.09, 131.95, 131.77, 131.50,
5
–1
hol. The yield 0.137 g (45%), mp 213 °C; IR: 3360–2800 cm
131.42, 130.85, 130.67, 130.07, 129.97 (C-Ph), 122.65 (C4),
39.99 (CH ).Anal. calcd. for C H N O : C, 70.81; H, 4.91;
–
1
–1
(
b, OH, COOH), 1668 cm (C=O, COOH), 1650 cm (C=O,
3
18 15 3 2
1
benzoyl); H-NMR (CDCl ): d = 10.18–10.11 (b, 1H, OH),
.67–7.13 (m, 10H, Harom), 3.90 ppm (s, 3H, CH3); C-
N, 13.77. Found: C, 70.83; H, 4.88; N, 13.75.
3
1
3
7
NMR (CDCl ): d = 195.47 (C=O, benzoyl), 164.52 (C=O,
3.5. Butyl-4-benzoyl-1-methyl-5-phenyl-1H-pyrazole-3-
3
COOH), 149.09 (C ), 144.27 (C ), 139.18, 134.98, 131.93,
carboxylate 5b
3
5
1
4
31.79, 131.34, 130.70, 130.02, 129.49 (C-Ph), 122.63 (C4),
0.40 ppm (N-CH ). Anal. calcd. for C H N O : C, 70.58;
The acid chloride 4 (0.324 g, 1 mmole) and excess of
n-butyl alcohol (molar ratio 1:3) were refluxed together with
catalytic amounts of pyridine for 2–3 h.After cooling the solu-
tion was acidified by adding diluted hydrochloric acid to give
a crude solid, recrystallized from n-butyl alcohol. The yield
was 0.159 g (44%), mp 99–100 °C; IR: 1703 (C=O, ester),
3
18 14 2 3
H, 4.61; N, 9.15. Found: C, 70.65; H, 4.59; N, 9.14.
3.2. 5-Benzoyl-4-hydroxsy-2-methyl-6-phenyl-2H-
pyridazin-3-one 3
–
1
1
After solvent of the filtrate of 2 was removed by evapora-
tion, the oily residue treated with ether and the formed crude
product was recrystallized from acetic acid to give 0.122 g
1664 cm (C=O, benzoyl). H-NMR (CDCl ): d = 7.78–
3
7.26 (m, 10H, Harom), 4.06–4.00 (t, 2H, OCH ), 3.90 (s, 3H,
2
N–CH ), 1.33–1.02 (m, 4H, –CH –CH –), 0.79-0.72 (t, 3H,
3
2
2
–
1
–1
13
(
40%) of 3, mp 218 °C; IR: 3117 cm (OH), 3080 cm (Ar-
–CH3), C-NMR (CDCl ): d = 193.12 (C=O, benzoyl),
3
–
1
–1
1
H), 1669 cm (C=O, benzoyl), 1603 cm (C=O); H-NMR
163.41 (C=O, ester), 146.36, (C ), 143.01 (C ), 140.23,
3
5
(
3
CDCl ): d = 9.41 (b, H, OH), d = 7.8–7.2 (m, 10H, Harom),
135.09, 131.64, 131.31, 130.76, 130.35, 129.67, (C-Ph),
124.09 (C ), 67.05 (O–CH ), 40.73 (N-CH ), 32.19, 20.89
3
13
.9 ppm (s, 3H, CH3); C-NMR (CDCl ): d = 193.56 (C=O,
3
4
2
3
benzoyl), 159.21 (C=O), 152.34 (C ), 148.88 (C ), 138.20,
(–CH –), 15.58 (CH ).Anal. calcd. for C H N O : C, 72.91;
4
5
2 3 22 22 2 3
1
37.11, 136.15, 131.52, 131.22, 130.81, 130.46, 130.27,
H, 6.12; N, 7.73. Found: C, 72.88; H, 6.08; N, 7.72.

E. Akbas et al. / IL FARMACO 60 (2005) 23–26
25
3
.6. N-[4-Benzoyl-1-methyl-5-phenyl-1H-pyrazole-3-car-
described by Clause [9] with minor modifications. Ampicil-
lin was used as standard antibacterial agent. Solutions of the
test compounds and ampicillin were dissolved in DMSO at
bonyl]-N′-phenyl urea 5c
–
1
Compound 4 (0.324 g, 1 mmole) and phenyl urea (0.136 g,
mmole) after refluxing in xylene for 6 h and cooling to
concentration of 5000 µg ml . The twofold dilution of the
compounds and ampicillin were prepared (2500, 1250, 620,
1
–1
room temperature the precipitate was filtered off and recrys-
tallized from xylene, yielding 0.212 g (50%), mp 227 °C; IR:
3
310, 150, 70...) µg ml . MIC tests were carried out in Muller-
Hinton Broth (Difco) medium, pH 7.2, with an inoculum of
–
1
–1
–1
6
–1
–1
244–3145 cm (NH), 1711 cm (C=O, urea), 1674 cm
(1–2) × 10 Colony Forming Unit ml (CFU ml ) four
microorganisms. The chemical compounds Muller-Hinton
Broth serial tube dilutions inoculated with each bacterium
were incubated on a rotary shaker at 37 °C for 18 h at 150 rpm.
The minimum inhibitory concentrations of the chemical com-
pounds were recorded as the lowest concentration of each
chemical compounds in the tubes with no growth (i.e., no
turbidity) of inoculated four bacteria. Muller-Hinton Broth
medium containing DMSO inoculated microorganisms were
used as a control. Experiments were repeated at least three to
find out reproducibility of the assays.
–
1
1
(C=O), 1661 cm (C=O). H-NMR (CDCl ): d = 10.24 (s,
3
1H, NH), 9.14 (s, 1H, NH), 7.81–7.04 (m, 15H, Harom), 3.92
1
3
(s, 3H, CH3). C-NMR (CDCl ): d = 193.19 (C=O), 163.14
3
(C=O), 152.23 (C=O, urea), 147.32 (C ), 143.32 (C ), 139.44,
3
5
1
1
39.22, 135.46, 131.88, 131.50, 130.90, 130.43, 129.26,
26.18, 123.90 (C-Ph), 122.35 (C ), 40.30 (CH ).Anal. calcd.
4
3
for C H N O : C, 70.74 ,H, 4.75; N, 13.20. Found: C, 70.75;
H, 4.73; N, 13.21.
25 20 4 3
3
.7. 4-Benzoyl-1-methyl-5-phenyl-1H-pyrazole-3-carboni-
trile 6
A cold solution of the acid amide 5a (0.287 g, 1 mmole) in
5. Statistical analysis
a mixture of DMF (0.7 ml) and SOCl (0.15 ml) was stirred
2
at 0–5 °C for 2 h. After heating to room temperature, stirring
was continued for overnight, then the reaction mixture poured
over crushed ice and the separated solid filtered off, washed
with water and recrystallized from methanol to give 0.186 g
Analysis of variance (ANOVA) was performed using a gen-
eral linear model procedure. A randomized complete block
design with repeated measurements was applied in this study.
The significance of differences of the MIC values of three
replicates were determined at the 95% confidence limit
(P = 0.05). Differences among treatments were examined for
levels of significance by the least significant differences (LSD)
test (SAS Inst., Inc., Cary, NC).
–1
–1
(
(
65%) of 6, mp: 169 °C; IR: 2237 cm (CN), 1676 cm
1
3
C=O, benzoyl). C-NMR (CDCl -d ): d = 192.28 (C=O,
3
6
benzoyl), 162.69 (C ), 144.36 (C ), 141.53 (C ), 140.42,
3
5
4
1
1
39.22, 136.07, 133.85, 133.50, 133.20, 133.08, 132.74,
14.13 (C≡N), 41.72 ppm (CH ).Anal. calcd. for C H N O:
3
18 13 3
C, 75.26; H, 4.53; N, 14.63. Found: C, 75.33; H, 4.52; N,
14.64.
6. Results and discussion
In this study, compounds 2, 3, 5a–c and 6 have been evalu-
4. Antibacterial Activity
ated for their antibacterial activity against Gram-negative
(
Escherichia coli ATCC 4230 and Pseudomonas putida
The chemical compounds were tested for antibacterial
ATCC) and Gram-positive (Bacillus cereus ATCC 7064 and
–
1
activity against human pathogenic Gram-negative (E. coli
ATCC 4230, Pseudomonas putida) and Gram-positive bac-
teria (Staphylococcus aureus ATCC 6538, Bacillus cereus
ATCC 7064). The minimal inhibitory concentrations (MICs)
of the chemical compounds assays were carried out as
Staphylococcus aureus ATCC 6538). The MICs (µg ml ) of
tested compounds against bacteria are showed in Table 1.
The results revealed that all chemical compounds exhib-
ited promising antibacterial activity against test bacteria. Com-
pounds had relatively a good antibacterial activity against
Table 1
a
b
Minimal inhibitory concentrations (MICs) of the compounds 2, 3, 5a–c and 6 derivatives against Gram–positive and Gram–negative bacteria
Compound
Bacillus cereus
ATCC 7064
1250
Staphylococcus aureus
ATCC 6538
620
Escherichia coli
ATCC 4230
620
Pseudomonas putida
2
3
1250
620
620
620
310
310
25
1250
620
620
5
5
5
6
a
b
c
620
310
620
620
620
310
620
310
310
620
620
620
Ampicillin
50
12
12
a
–1
MICs values were determined as µg ml active compounds in medium.
b
–1
The MIC values (µg ml ) with different letters in the same column are significantly different (P < 0.05).

26
E. Akbas et al. / IL FARMACO 60 (2005) 23–26
Gram-negative bacteria more than Gram-positive bacteria.
Particularly, the compound named as 5c exhibited more anti-
bacterial potencies than other compounds against Gram-
positive and Gram-negative bacteria (Table 1). In addition,
did not found significantly differences between antibacterial
activities of other compounds against test microorganisms.
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