Synthesis and biological evaluation of novel pyrazole derivatives as antibacterial agents

Article abstract of DOI:10.3184/174751912X13450480267670

The synthesis of a novel series of 5-[(X-benzoylamino)]-1-(2,4- dinitrophenyl)-1H-4-pyrazolecarboxamide derivatives is described. Their antibacterial activity (MIC) against methicillin-susceptible and methicillin-resistant Staphylococcus aureus (MRSA) wer

Full text of DOI:10.3184/174751912X13450480267670

JOURNAL OF CHEMICAL RESEARCH 2012
RESEARCH PAPER 579
OCTOBER, 579–580
Synthesis and biological evaluation of novel pyrazole derivatives as
antibacterial agents
Fariba Daemi, Sadegh Allameh and Mehdi Pordel*
Department of Chemistry, Mashhad Branch, Islamic Azad University, Mashhad, Iran
The synthesis of a novel series of 5-[(X-benzoylamino)]-1-(2,4-dinitrophenyl)-1H-4-pyrazolecarboxamide derivatives
is described. Their antibacterial activity (MIC) against methicillin-susceptible and methicillin-resistant Staphylococ-
cus aureus (MRSA) were determined. The results revealed that two of the compounds, X = 4-MeO and X = 4-NO2,
displayed greater antibacterial activity against MRSA than did cephalexin, cloxacillin and erythromycin.
Keywords: antibacterial agents, pyrazoles, Staphylococcus aureus, MRSA, MIC
Resistance to antimicrobial agents is recognised as a major
global public health problem, so that the discovery of new anti-
bacterial and antifungal compounds has become increasingly
critical in fighting infectious disease.
In continuation of our previous studies on the activity of
trisubstituted pyrazoles against bacteria,1,2 we report the syn-
thesis of some analogues of those compounds which show
much improved antimicrobial activities against gram positive
bacteria.
These compounds were effective only against gram positive
bacteria, and their activities against MRSA and MSSA are
shown in Table 1. Synthetic intermediates 1 and 2 were not
effective at all. Compounds 4a–d and 4f which have para
substitutents in the R group, showed the best inhibitory effects
against both MRSA and MSSA. 4e, 4g and 4h were found to
exhibit moderate antibacterial activities. Gratifyingly, these
results clearly show that when the cyano group in compound 1
is hydrolysed to the corresponding amide 2, and the latter
converted to new compounds 4a–h, they exhibited powerful
antibacterial activity in comparison with previous pyrazole
derivatives which we have synthesised.1,2 As the data in Table
1 show, compound 4f shows greater inhibitory activity against
MRSA and MSSA than did the well known antibacterial agents
cephalexin, cloxacillin and erythromycin.
To summarise, we have synthesised some novel trisubsti-
tuted pyrazoles and shown them to be very effective S. aureus
growth inhibitors. Such compounds would appear to offer a
suitable template for the design of more powerful antibacterial
agents and further studies are under way to this end in our
laboratory.
Results and discussion
We have reported1,2 the synthesis and biological evaluation of
a series of N-acylated derivatives of 5-amino-1-(2,4-dinitro-
phenyl)-1H-4-pyrazolecarbonitrile 1 (see Scheme 1). Here we
report on the synthesis and testing of the corresponding N-acyl
derivatives of the amido compound 2.
Compound 1, which was readily obtained1,2 from the reac-
tion of 2,4-dinitrophenylhydrazine with 2-(ethoxymethylene)
malononitrile in ethanol under reflux using a literature method,3
was smoothly hydrolysed by sulfuric acid at room tempera-
ture4 to afford 5-amino-1-(2,4-dinitrophenyl)-1H-4-pyrazole-
carboxamide 2. The new 5-acylamino-1-(2,4-dinitrophenyl)-
1H-4-pyrazolecarboxamides 4a–h were synthesised from the
reaction of compound 2 and the corresponding acid chlorides
3a–h in dry pyridine (Scheme 1).
Experimental
Melting points were recorded on an Electrothermal type-9100 melt-
ing-point apparatus. The IR spectra (as KBr discs) were obtained on a
Tensor 27 spectrometer. 1H NMR spectra were recorded at 400 Hz on
a Bruker Avance DRX-400 FT spectrometer in DMSO-d6. Chemical
shifts are reported in ppm downfield from TMS as internal standard;
coupling constants J are given in Hz. The mass spectra were recorded
on a Varian Mat, CH-7 at 70 eV. Elemental analysis was performed on
a Thermo Finnigan Flash EA microanalyser.
All chemicals were purchased from Sigma, Fluka or Merck Co.
S. aureus ATCC 1112 was purchased from the Pasteur Institute of
Iran and S. aureus (methicillin-resistant) was isolated from different
specimens which were referred to the Microbiological Laboratory,
Ghaem Hospital, Medical University of Mashhad-Iran. Its methicillin
resistance was tested according to the NCCLS guidelines.5
Structural assignments of compounds 4a–h were based
upon their spectral and microanalytical (C, H and N) data.
The synthetic compounds 1, 2 and 4a–h were screened for
antibacterial activity against Escherichia coli HB101 (BA-
7601C), Staphylococcus aureus pathogens (methicillin-resis-
tant S. aureus (MRSA) and methicillin-susceptible S. aureus
(MSSA - ATCC 1112)), Pseudomonas aeruginosa (PTCC
1431), and Bacillus subtilis (PTCC 1365). Experimental details
of the tests can be found in our earlier study.1
Scheme 1
* Correspondent. E-mail: mehdipordel58@yahoo.com

580 JOURNAL OF CHEMICAL RESEARCH 2012
Table 1 Antibacterial activity (MIC, µg mL−1) of 5-[(X-benzoyla
mino)]-1-(2,4-dinitrophenyl)-1H-4-pyrazolecarboxamides 4a–h
against MRSA and MSSA
(430.76): C, 47.40; H, 2.57; N, 19.51. Found: C, 47.23; H, 2.49; N,
19.65%.
5-[(4-Bromobenzoyl)amino]-1-(2,4-dinitrophenyl)-1H-4-pyrazole-
carboxamide (4d): Light yellow crystals (ethanol). Yield, 71%; m.p.
299–301 °C; IR: 1690, 1703 (C=O), 3217 (NH) and 3185, 3390 (NH2)
cm−1; 1H NMR: δ = 7.77 (d, 2H, J = 8.3), 7.87 (d, 2H, J = 8.3), 8.17
(d, 1H, J = 8.8 Hz), 8.33 (s, 1H), 8.77 (dd, 1H, J = 8.8 Hz, J′ = 2.4 Hz),
8.85 (br s, 1H), 8.92 (d, 1H, J = 2.4 Hz), 8.97 (br s, 1H), 10.50 (br s,
1H); MS (70 eV): m/z = 477 (M++2). Anal. Calcd for C17H11N6O6Br
(475.21): C, 42.97; H, 2.33; N, 17.68. Found: C, 42.73; H, 2.23; N,
17.91%.
Compd MRSA MSSA
Compd
MRSA MSSA
1 and 2
4a
–
–
4f
4g
4.0
75.3
99.4
32.0
94.0
72.0
4.0
75.9
99.4
32.0
13.7
4.6
10.0
42.0
39.0
29.4
45.3
10.0
42.0
35.3
29.4
45.3
4b
4h
4c
Erythromycin
Cloxacillin
Cephalexin
4d
4e
5-[(3-Chlorobenzoyl)amino]-1-(2,4-dinitrophenyl)-1H-4-pyrazole-
carboxamide (4e): Light yellow crystals (ethanol). Yield, 55%; m.p.
251–253 °C; IR: 1690, 1705 (C=O), 3221 (NH) and 3185, 3390 (NH2)
cm−1; 1H NMR: δ = 7.35–7.39 (m, 1H), 7.54 (1H, t, J = 7.9 Hz), 7.74–
7.80 (m, 2H), 7.95 (d, 1H, J = 8.8 Hz), 8.14 (s, 1H), 8.56 (dd, 1H,
J = 8.8 Hz, J′ = 2.4 Hz), 8.79 (d, 1H, J = 2.4 Hz), 8.87 (br s, 1H), 8.99
(br s, 1H), 9.30 (br s, 1H); MS (70 eV): m/z = 432 (M++2). Anal. Calcd
for C17H11N6O6Cl (430.76): 47.40; H, 2.57; N, 19.51. Found: C, 47.21;
H, 2.50; N, 19.31%.
1-(2,4-Dinitrophenyl)-5-[(4-nitrobenzoyl)amino]-1H-4-pyrazole-
carboxamide (4f): Yellow crystals (ethanol). Yield, 60%; m.p. 306–
307 °C; IR: 1690, 1706 (C=O), 3220 (NH) and 3185, 3390 (NH2)
cm−1; 1H NMR: δ = 7.69 (d, 2H, J = 8.0 Hz), 7.78 (d, 2H, J = 8.0 Hz),
8.22 (d, 1H, J = 8.8 Hz), 8.29 (s,1H), 8.75 (dd, 1H, J = 8.8 Hz, J´ =
2.4 Hz), 8.85 (d, 1H, J = 2.4 Hz), 8.89 (br s, 1H), 9.05 (br s, 1H), 10.29
(br s, 1H); MS (70 eV): m/z = 441 (M+). Anal. Calcd for C17H11N7O8
(441.31): C, 46.27; H, 2.51; N, 22.22. Found: C, 46.03; H, 2.43; N,
22.34%.
MRSA, methicillin-resistant Staphylococcus aureus,
MSSA, methicillin-susceptible Staphylococcus aureus.
Synthesis of 5-[(x-benzoylamino)]-1-(2,4-dinitrophenyl)-1H-4-
pyrazolecarboxamides 4a–h; general procedure
A mixture of ethoxymethylenmalononitrile (12.2 g, 100 mmol) and
2,4-dinitrophenylhydrazine (19.8 g, 100 mmol) in ethanol (200 mL)
was heated under reflux for 4 h. After cooling, the crystals that formed
were filtered off, washed with ethanol and dried at 70 °C to give
compound 1 (18.4 g, 67%, m.p. 217–220 °C), [lit.3 218–220 °C].
Compound 1 (34 mmol) was gradually added during 1 h, with
stirring to concentrated sulfuric acid (30 mL), which was kept in an
ice-bath. The inside temperature was kept between 15 and 20 °C. The
solution was stirred at rt for a further 4 h, then water (40 mL) was
added to the solution in an ice-bath and it was stirred for a further 2 h
before it was poured onto crushed ice; finally it was neutralised with
dilute aqueous NaOH. The reaction mixture which was allowed to
reach 50–70 °C during the neutralisation, was cooled to rt, filtered and
washed with water and then CH2Cl2, to give the pure product 2 (9.2 g,
93%, m.p. 246–247 °C), [lit.4 246–248 °C].
5-(Benzoylamino)-1-(2,4-dinitrophenyl)-1H-4-pyrazolecarboxamide
(4g): Light yellow crystals (ethanol). Yield, 70%; m.p. 239–240 °C;
1
IR: 1690, 1704(C=O), 3225 (NH) and 3185, 3390 (NH2) cm−1; H
Acid chloride (3a–h) (12 mmol) was added dropwise at room
temperature to a stirred solution of 2 (2.74 g, 10 mmol) in dry pyridine
(20 mL). After refluxing for 4 h, the pyridine was evaporated under
reduced pressure. The residue was treated with 5% sodium carbonate
(2 × 50 mL) and extracted with dichloromethane (2 × 30 mL). The
organic extract was dried with anhydrous sodium sulfate, concentrated
under reduced pressure and crystallised to provide the pure desired
compound 4a–h.
NMR: δ = 7.35–7.65 (m, 5H), 8.05 (d, 1H, J = 8.8 Hz), 8.11 (s, 1H),
8.28 (br s, 1H), 8.65 (dd, 1H, J = 8.8 Hz, J´ = 2.4 Hz), 8.88 (d, 1H,
J = 2.4 Hz), 8.97 (br s, 1H), 9.09 (br s, 1H); MS (70 eV): m/z = 396
(M+). Anal. Calcd for C17H12N6O6 (396.32): C, 51.52; H, 3.05; N,
21.21. Found: C, 51.35; H, 2.95; N, 21.01%.
5-(Acetylamino)-1-(2,4-dinitrophenyl)-1H-4-pyrazolecarboxamide
(4h): Light yellow crystal (ethanol).Yield, 77%; m.p. 320–321 °C; IR:
1690, 1695(C=O), 3221 (NH) and 3185, 3390 (NH2) cm−1; 1H NMR:
δ = 2.01 (s, 3H), 8.05 (d, 1H, J = 8.8 Hz), 8.35 (s, 1H), 8.46 (dd, 1H,
J = 8.8 Hz, J´ = 2.4 Hz), 8.73 (d, 1H, J = 2.4 Hz), 8.87 (br s, 1H), 8.94
(br s, 1H), 10.67 (br s, 1H); MS (70 eV): m/z = 334 (M+). Anal. Calcd
for C12H10N6O6 (334.25): C, 43.12; H, 3.02; N, 25.14. Found: C, 42.90;
H, 2.93; N, 25.40%.
1-(2,4-Dinitrophenyl)-5-[(4-methoxybenzoyl)amino]-1H-4-pyra-
zolecarboxamide (4a): Light yellow crystals (ethanol). Yield, 69%;
m.p. 275–277 °C; IR: 1667, 1690 (C=O), 3243 (NH) and 3183, 3395
1
(NH2) cm−1; H NMR: δ = 3.86 (s, 3H), 6.97 (d, 2H, J = 8.5), 7.77
(d, 2H, J = 8.5), 8.13 (d, 1H, J = 8.8 Hz), 8.17 (s, 1H), 8.45 (br s, 1H),
8.60 (dd, 1H, J = 8.8 Hz, J′ = 2.4 Hz), 8.73 (br s, 1H), 8.88 (br s, 1H),
8.95 (d, 1H, J = 2.4 Hz); MS (70 eV): m/z = 426 (M+). Anal. Calcd for
C18H14N6O7 (426.34): C, 50.71; H, 3.31; N, 19.71. Found: C, 50.59; H,
3.25; N, 20.01%.
1-(2,4-Dinitrophenyl)-5-[(4-methylbenzoyl)amino]-1H-4-pyra-
zolecarboxamide (4b): Light yellow crystals (ethanol). Yield, 73%;
m.p. 260–262 °C; IR: 1667, 1690 (C=O), 3228 (NH) and 3185, 3390
(NH2) cm−1; 1H NMR: δ = 2.43 (s, 3H), 7.43 (d, 2H, J = 8.3), 7.75 (d,
2H, J = 8.3), 8.27 (d, 1H, J = 8.8 Hz), 8.31 (s, 1H), 8.55 (dd, 1H, J =
8.8 Hz, J′ = 2.4 Hz), 8.77 (br s, 1H), 8.95 (br s, 1H), 8.99 (d, 1H, J =
2.4 Hz), 9.45 (br s, 1H); MS (70 eV): m/z = 410 (M+). Anal. Calcd for
C18H14N6O6 (410.34): C, 52.69; H, 3.44; N, 20.48. Found: C, 52.49; H,
3.37; N, 20.65%.
5-[(4-Chlorobenzoyl)amino]-1-(2,4-dinitrophenyl)-1H-4-pyrazole-
carboxamide (4c): Light yellow crystals (ethanol). Yield, 65%; m.p.
284–285 °C; IR: 1690, 1704 (C=O), 3217 (NH) and 3185, 3390 (NH2)
cm−1; 1H NMR: δ = 7.48 (d, 2H, J = 8.4), 7.73 (d, 2H, J = 8.4), 8.07
(d, 1H, J = 8.8 Hz), 8.23 (s, 1H), 8.67 (dd, 1H, J = 8.8 Hz, J′ = 2.4 Hz),
8.73 (br s, 1H), 8.85 (br s, 1H), 8.91 (d, 1H, J = 2.4 Hz), 9.12 (br s,
1H); MS (70 eV): m/z = 432 (M++2). Anal. Calcd for C17H11N6O6Cl
We would like to express our sincere gratitude to Dr Hamid
Sadeghian (Mashhad University of Medical Sciences,
Mashhad, Iran) for financial support of this work.
Received 28 June 2012; accepted 3 July 2012
Paper 1201371 doi: 10.3184/174751912X13450480267670
Published online: 28 September 2012
References
1
M. Rahimizadeh, M. Pordel, M. Bakavoli, Sh. Rezaeian and A. Sadeghian,
World. J. Microbiol. Biotechnol., 2010, 26, 317.
2
H. Sadeghian, A. Sadeghian, M. Pordel, M. Rahimizadeh, P. Jahandari,
A. Orafaie and M. Bakavoli, Med. Chem. Res., 2010, 19, 103.
3
4
A. Kreutzberger and K. Burgwitz, J. Heterocycl. Chem., 1980, 17, 265.
M. Pordel, Synthesis of new derivatives of aminopyrazolocarbonitrile,
2004, MSc thesis, Ferdowsi, University of Mashhad, Mashhad, Iran.
S.M. Finegold and L. Garrod, Bailey and Scott’s diagnostic microbiology,
C.V. Mosby Co., Toronto, 1995, 3rd edn., Chap. 13, pp. 171–193.
5

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