Synthesis and biological testing of novel analogues of sydnone as potential antibacterial agents.

Article abstract of DOI:10.1002/ardp.200300814

Several series of 3-phenylsydnone derivatives conjugated to well-known moieties with antibacterial activity were synthesized via several routes. These derivatives include 3-cyano-2-oxopyridine, 2-amino-3-cyanopyridine, 2-arylidene-1-ethylidenehydrazine and 2-aroyl-1-ethylidenehydrazine moieties. Thus, the key intermediate 3-(4-acetylphenyl)sydnone (3) was allowed to react with the appropriate aldehyde, ethyl cyanoacetate or malononitrile in presence of excess ammonium acetate in two steps (method 1) or through a one-pot reaction technique (methods 2 and 3) to give the corresponding sydnone derivatives 5 and 6, respectively. Moreover, condensation of compound 3 with hydrazine hydrate followed by the reaction with the appropriate aldehyde, mono- and dicarboxylic acid hydrazide yielded the corresponding sydnone derivatives 8, 9 and 10, respectively. Most of the synthesized compounds were screened for their in vitro antibacterial activity against various pathogenic organisms of both Gram-positive and Gram-negative bacteria. The minimum inhibitory concentrations (MICs) were determined using agar dilution method.

Full text of DOI:10.1002/ardp.200300814

164 Nasr et al.
Arch. Pharm. Pharm. Med. Chem. 2004, 337, 164−170
Mohamed A. Moustafaa,
Magda N. Nasrb,
Synthesis and Biological Testing of Novel
Analogues of Sydnone as Potential
Antibacterial Agents
Magdy M. Gineinahb,
Waleed A. Bayoumib
a Department of
Several series of 3-phenylsydnone derivatives conjugated to well-known moiet-
ies with antibacterial activity were synthesized via several routes. These deriva-
tives include 3-cyano-2-oxopyridine, 2-amino-3-cyanopyridine, 2-arylidene-1-
ethylidenehydrazine and 2-aroyl-1-ethylidenehydrazine moieties. Thus, the key
intermediate 3-(4-acetylphenyl)sydnone (3) was allowed to react with the appro-
priate aldehyde, ethyl cyanoacetate or malononitrile in presence of excess am-
monium acetate in two steps (method 1) or through a one-pot reaction technique
(methods 2 and 3) to give the corresponding sydnone derivatives 5 and 6,
respectively. Moreover, condensation of compound 3 with hydrazine hydrate fol-
lowed by the reaction with the appropriate aldehyde, mono- and dicarboxylic
acid hydrazide yielded the corresponding sydnone derivatives 8, 9 and 10,
respectively. Most of the synthesized compounds were screened for their in vitro
antibacterial activity against various pathogenic organisms of both Gram-posi-
tive and Gram-negative bacteria. The minimum inhibitory concentrations (MICs)
were determined using agar dilution method.
Pharmaceutical Chemistry,
Faculty of Pharmacy, King
Abdulazis University,
Geddah, Saudi Arabi
b Department of
Pharmaceutical Organic
Chemistry, College of
Pharmacy, University of
Mansoura, Mansoura, Egypt
Keywords: Sydnone; Cyanooxopyridine; Aminocyanopyridine; Arylidene-
hydrazine; Aroylhydrazine; Antibacterial agents
Received: April 28, 2003; Accepted: October 14, 2003 [FP814]
DOI 10.1002/ardp.200300814
Introduction
bacterial activity [11, 2]. The biological activity associ-
ated with these compounds was attributed to the pres-
Research on the field of antimicrobial agents is of sig-
nificant interest in the scope of medicinal chemistry
aiming for the discovery of new agents having poten-
tial activity, broader spectrum, less toxicity, and safer
therapeutic profiles than the currently available ones.
The primary aim was to synthesize different sydnones
conjugated with effective and well-known pharmac-
ophoric moieties of potential antibacterial activity, as
for example, cyanooxopyridine, aminocyanopyridine
and substituted hydrazine moieties. Several publi-
cations have pointed out the value of sydnones as
antimicrobial agents [1Ϫ5], as compounds IϪIV (Fig-
ure 1). It was observed that in the majority of the re-
ported compounds, the presence of substituted phenyl
group at the 3-position of the sydnone was a common
structural feature. On the other hand, the antimicrobial
activity of pyridine ring substituted with a wide variety
of functionalities was reported [6Ϫ8]. Cyanooxopyrid-
ines and aminocyanopyridines were among pyridine
derivatives that show significant activity [9, 10]. In ad-
dition, earlier reports revealed that several hydrazones
e.g., verazide V, ftivazide VI and others, exhibit anti-
ence of the (ϪCONHϪN=CHϪ) moiety. Likewise, sev-
eral hydrazine derivatives especially 2-arylidene and
2-aroylhydrazines were reported to possess good anti-
bacterial activity, e.g. isonicotinyl hydrazones [13] and
compounds with the general structure VII [14] (Figure
1). Taking the above mentioned considerations into
account, molecular conjugation of the sydnone moiety
with two or more active counterparts was designed
and synthesized with the hope of producing novel
sydnone derivatives that possess good antibacterial
activity.
Results and discussion
Chemistry
The synthetic strategies adopted to obtain the target
compounds are summarized in Schemes 1-3. The
standard procedure that was reported earlier [15, 16]
and later by Baker et al. [17Ϫ19] was adopted for the
preparation of the compound 3-(4-acetylphenyl)syd-
none (3) (Scheme 1). Reaction of equimolar amounts
of 4-aminoacetophenone with aqueous solution of so-
dium salt of chloroacetic acid yielded N-(4-acetylphe-
nyl)glycine (1), which was nitrosated by the action of
sodium nitrite in dilute hydrochloric acid solution to
Correspondence: Magda N. Nasr, Department of Pharma-
ceutical Organic Chemistry, College of Pharmacy, University of
Mansoura, Mansoura 35516, Egypt. Phone: +20 50 224-7496,
Fax: +20 50 224-7496; e-mail: magdaahnasr@yahoo.com
 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Arch. Pharm. Pharm. Med. Chem. 2004, 337, 164−170
Novel analogues of sydnone as potential antibacterial agents 165
The first one was applied through the formation of the
intermediates 3-[4-(3-aryl-1-oxo-2-propen-1-yl)phenyl-
]sydnones 4aϪg via Claisen-Schmidt condensation of
3-(4-acetylphenyl)sydnone (3), which was sub-
sequently allowed to react with the appropriate aro-
matic aldehyde in alcoholic sodium hydroxide (method
1) [3, 22]. Moreover, it was reported that cyanooxopyri-
dines were synthesized in a one-pot reaction by heat-
ing compound 3 with the aldehydes in presence of am-
monium acetate neat [23] or in a suitable solvent [24-
26]. In the present work, compounds 5aϪj were syn-
thesized by condensation of compound 3 with the ap-
propriate aromatic aldehyde and ethyl cyanoacetate in
presence of excess ammonium acetate using ethanol
as solvent through a one-pot reaction (method 2) or
by heating the previous reaction mixture neat in an oil
bath at 140-160°C (method 3). All members of the
series 5aϪj were prepared by the 3 methods except
compounds 5c, 5e and 5i which were prepared by
methods 2 and 3 only. Comparison of the yield data
of the 3 methods revealed that the one-pot reaction
methods were the best in terms of yield percentage
especially when the reaction was carried out neat
(method 3) (Table 1).
Figure 1. Representative examples of sydnones and
hydrazine derivatives of antimicrobial acitivities.
Construction of an aminocyanopyridine ring could be
achieved by similar methods [27Ϫ30] applied for the
synthesis of cyanooxopyridines except using malonon-
itrile instead of ethyl cyanoacetate. Thus, sydnone
analogues 6aϪf were synthesized by the reaction of
compounds 4aϪg, malononitrile and ammonium acet-
ate in ethanol or n-butanol as solvents (method 1).
They had been also synthesized by condensation of
Scheme 1.
give the N-nitroso-N-(4-acetylphenyl)glycine (2). The
N-substituted N-nitrosoglycines were reported to be
cyclized to the corresponding sydnones by the action
of trifluoroacetic anhydride in dichloromethane at room
temperature [18], or by heating in acetic anhydride [20,
21]. Compound 2 was successfully cyclized to its syd-
none analogue 3 in a 75% yield by the action of acetic
anhydride (Scheme1) [3]. Compound 3 was con-
sidered an important synthon for the construction of
the pyridine ring. 3-[4-(4-Aryl-3-cyano-1,2-dihydro-2-
oxo-6-pyridyl)phenyl]sydnones 5aϪj were synthesized
following the two strategies illustrated in Scheme 2.
Scheme 2.
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166 Nasr et al.
Arch. Pharm. Pharm. Med. Chem. 2004, 337, 164−170
Table 1. Physicochemical data of the new compounds.
Comp.
Ar
MP (°C)
Solvents†
Yield (%)
A/B/C‡
Mol. Formula
5a
5b
5c
5d
5e
5f
5g
5h
5i
C6H5
268Ϫ270
256Ϫ257
280Ϫ282
273Ϫ275
160Ϫ162
178Ϫ180
252Ϫ254
>300
EA
EA
DW
DW
EA
EA
EA
DW
EE
DT
EA
DW
EA
DW
DW
DW
E
EA
E
ED
ED
ED
E
EA
DW
DW
DW
DW
DW
17/23/30
30/40/47
Ϫ/25/34
28/34/38
Ϫ/18/23
20/23/30
20/26/35
35/43/50
Ϫ/29/30
19/20/25
10/22/15
Ϫ/15/12
Ϫ/30/30
20/24/24
15/20/17
Ϫ/14/11
74
C20H12N4O3
C20H11BrN4O3
C20H11ClN4O3
C20H11ClN4O3
C22H17N5O3
C20H12N4O4
C21H14N4O4
C20H11N5O5
C20H10ClN5O5
C18H10N4O4
C20H13N5O2
C20H12BrN5O2
C20H12BrN5O2
C20H12ClN5O2
C21H15N5O3
C20H12N6O4
C17H14N4O2
C17H13BrN4O2
C17H13ClN4O2
C17H14N4O3
C17H13N5O4
C17H12ClN5O24
C17H12Cl2N4O2
C15H12N4O3
C17H14N4O4
C17H14N4O4
C16H13N5O3
C15H12N4O4
C23H20N8O6
2-Br-C6H4
2-Cl-C6H4
4-Cl-C6H4
4-(CH3)2N-C6H4
4-HO-C6H4
4-H3CO-C6H4
4-NO2-C6H4
2-Cl-5-NO2-C6H3
2-Furyl
191Ϫ193
>300
5j
6a
6b
6c
6d
6e
6f
8a
8b
8c
8d
8e
8f
8g
8h
9a
9b
9c
9d
10
C6H5
193Ϫ195
205Ϫ207
187Ϫ189
239Ϫ241
263Ϫ265
246Ϫ248
173Ϫ175
184Ϫ185
189Ϫ190
196Ϫ197
192Ϫ193
135Ϫ236
186Ϫ188
167Ϫ168
225Ϫ227
258Ϫ260
234Ϫ236
205Ϫ207
210Ϫ212
2-Br-C6H4
4-Br-C6H4
4-Cl-C6H4
4-H3CO-C6H4
4-NO2-C6H4
C6H5
4-Br-C6H4
4-Cl-C6H4
4-HO-C6H4
4-NO2-C6H4
2-Cl-5-NO2-C6H3
2,6-Cl2-C6H3
2-Furyl
55
67
55
72
60
83
42
73
60
68
70
65
C6H5
4-HO-C6H4
4-Pyridyl
2-Furyl
Ϫ
†
Solvents: DT: DMF/toluene; DW: DMF/water; E: ethanol; EA: ethanol/acetone; ED: ethanol/ DMF; EE: ethanol/
ethyl acetate.
A: method 1, B: method 2, C: method 3.
‡
compound 3 malononitrile and the appropriate alde-
(4-acetylphenyl)sydnone (3) and hydrazine hydrate in
presence of ethanol as a solvent to yield compound
7 in a high yield. Condensation of the latter with the
appropriate aldehyde in ethanol gave the target com-
pounds 8a-h. Interaction of compound 3 with the ap-
propriate mono- and dicarboxylic acid hydrazide
yielded the corresponding sydnone derivatives 9aϪd
and 10, respectively (Scheme 3).
hyde in the presence of ammonium acetate using
ethanol as a solvent (method 2) or neat in an oil bath
at 140Ϫ160°C (method 3) through a one-pot reaction
technique (Scheme 2). All the synthesized compounds
belonging to the series 6aϪj were prepared by the 3
methods except compounds 6b, 6c and 6f were pre-
pared by methods 2 and 3 only. Comparison of the
yield data of the 3 methods showed that there was no
significant difference between them. On the other
hand, compound 3 was used as a precursor for the
synthesis of other target compounds 7Ϫ10. Com-
pounds 8aϪh were prepared by stirring at room tem-
perature for 8 h or refluxing for 1 h a mixture of 3-
Antibacterial Screening
We carried out the screening experiments in vitro for
antibacterial activities of the sydnone analogues. Four
 2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Arch. Pharm. Pharm. Med. Chem. 2004, 337, 164−170
Novel analogues of sydnone as potential antibacterial agents 167
of this series against both Gram-positive and Gram-
negative organisms and it is among the most active
compounds tested. However, compound 6a with an
unsubstituted phenyl ring was the least active of this
group of compounds. Alternatively, a nitrophenyl nu-
cleus in place of the phenyl ring gave compound 6f
with better activity against Gram-positive than Gram-
negative organisms. On the contrary, compound 7
manifested low antibacterial activity. On the other
hand, substitution of the N-2 of 7 with arylidene moiet-
ies gave compounds 8. Among this series of com-
pounds, compound 8b with a bromophenyl ring pos-
sessed the highest activities against all types of micro-
organisms. Against B. subtilis, 8b showed activity al-
most equal to that of the standard ciprofloxacin.
Compound 8h with a furyl group comes at the second
rank in activity against B. subtilis. Moreover, com-
pound 8c with a chlorophenyl moiety, is among the
most active members of the test compounds. Com-
pounds 9 with an aroyl group on the N-2 of compound
7 have moderate antibacterial activity with compound
9c being the least active. In summary, compounds 8b,
6c and 5b, having the common structural feature of a
bromophenyl moiety, exhibit the highest antibacterial
activities.
Scheme 3.
In conclusion, the results of the present study indicate
that sydnone derivatives containing pyridine nucleus
possess high antibacterial activity when a bromophe-
nyl group is incorporated into the pyridine nucleus. On
the other hand, replacement of the pyridine nucleus
with an unsubstituted hydrazone residue resulted in
formation of compound 7 with modest antibacterial ac-
tivity. However, substitution at the N-2 of the hydrazine
portion with an arylidene counterpart bearing a bromo
group resulted in compound 8b with the highest po-
tency and a broadened spectrum of antibacterial ac-
tivity in vitro. In addition, compounds 8c with a chloro-
benzylidene portion and 8h with a furylmethylidene
group as the arylidene part of this series showed high
activity against Gram-positive bacteria. Moreover, it is
worth noting that the replacement of the furylmethyl-
idene group in 8h with a furoyl moiety yielded
compound 9d which, unlike 8h, is of a moderate anti-
bacterial activity.
groups of sydnone were subjected to biological test-
ing. The first group comprises 3-phenylsydnone de-
rivatives with the phenyl ring bearing a pyridine nu-
cleus at the 4-position 5 and 6. The second structural
feature is represented by 7 with the 3-phenyl ring sub-
stituted at the 4-position with a hydrazone residue.
The remaining two groups have the N-2 of 7 substi-
tuted with either arylidene group 8 or aroyl functionality
9. To substantiate our antibacterial results, we scre-
ened these compounds against an assortment of two
Gram-positive and two Gram-negative organisms
using ciprofloxacin as a reference standard. The mini-
mum inhibitory concentrations (MICs, µg/mL) were de-
termined using standard agar dilution method [31].
The MIC values are summarized in Table 2. From the
obtained data, it is clear that all the tested members
of the pyridine series 5 and 6 possess higher activity
than the parent acetyl derivative 3. Among the pyridine
series, the lipophilic bromophenyl ring in compound 5b
showed high activity against both types of bacteria,
particularly, the Gram-negative organisms. On the
other hand, the less lipophilic hydroxyphenyl group of
5f revealed low activity while 5h with a nitrophenyl
residue possesses good activity. However, the order
of activity observed with the 2-oxopyridine derivatives
of sydnone 5 was also explored with the 2-aminopyrid-
ine series 6. It was found that compound 6c with
bromophenyl counterpart is the most active member
Acknowledgments
The authors deeply thank Dr. Samy M. Kheira, Depart-
ment of Microbiology, College of Pharmacy, Mansoura
University, Egypt, for carrying out the antibacterial
screening.
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168 Nasr et al.
Arch. Pharm. Pharm. Med. Chem. 2004, 337, 164−170
Table 2. Table 2. In vitro antibacterial activities of new compounds†,‡
Compound
Bacillus
subtilis
Staphylococcus
aureus
E. coli
Pseudomonas
aeruginosa
3
5b
5f
5h
6a
6c
6e
6f
64
16
64
32
64
8
32
16
64
16
1
>128
16
>128
32
64
8
16
16
>128
32
4
>128
8
64
8
64
16
>128
4
16
16
32
16
32
32
64
32
4
16
32
64
32
8
7
8a
8b
8c
8d
8e
8g
8h
9b
9c
9d
16
32
32
64
8
32
64
64
0.5
8
8
16
16
32
64
32
64
64
64
0.032
32
16
8
16
32
64
32
1
16
32
32
16
64
>128
32
0.064
Ciprofloxacin
†
MIC is the lowest concentration of compound needed for prevention of visible growth of microorganisms, re-
ported as the average values of duplicate determinations. MIC values obtained by the use of an agar dilution
method whereby organisms were deposited onto medicated agar plates by the replication device of Steers et
al. [31]
Organisms selected: Bacillus subtilis ATCC 6633; Staphylococcus aureus ATCC 29213; Escherichia coli ATCC
25922; Pseudomonas aeruginosa ATCIC 9027.
‡
rated solid was filtered, dried and recrystallized from the suit-
Experimental
able solvent. IR, 5a: 3450 (NH), 3144 (CH, sydnone), 2216
(CN), 1754 (C=O, sydnone), 1670 (C=O), 1580 (C=N). 5e:
3500 (NH), 3134 (CH, sydnone), 2210 (CN), 1759 (C=O, syd-
none), 1702 (C=O), 1609 (C=N). 5f: 3400 (OH), 3250 (NH),
3130 (CH, sydnone), 2213 (CN), 1745 (C=O, sydnone), 1656
(C=O), 1600 (C=N). 5h: 3350 (NH), 3103 (CH, sydnone),
2223 (CN), 1745 (C=O, sydnone), 1638 (C=O), 1585 (C=N).
5j: 3450 (NH), 3170 (CH, sydnone), 2216 (CN), 1754 (C=O,
sydnone), 1658 (C=O), 1580 (C=N). 1H-NMR (DMSO-d6): 5c:
7.28-8.40 (m, 10H, Ar-H, CH sydnone), NH seemed to be
exchanged by the solvent. 5d: 7.07-8.42 (m, 10H, Ar-H, CH
sydnone). 5g: 3.83 (s, 3H, OCH3), 7.00-8.40 (m, 10H, Ar-H,
CH sydnone), 12.85 (br s, 1H, NH; D2O exchangeable). 13C-
NMR (DMSO-d6): 5a: 168.4 (CO, sydnone), 162.3 (CO),
159.4, 149.9, 136.3, 135.8, 130.5, 129.5, 128.8, 128.3, 121.7
(Ar C), 116.2 (CN), 98.8 (C5, pyridine), 94.9 (CH, sydnone).
Chemistry
Melting points (°C, uncorrected) were determined on a
Fischer-Johns apparatus (Fischer-Scientific, USA). IR spec-
tra (KBr) were recorded on a Pye-Unicam SP1000 Spec-
trometer (Pye-Unicam Ltd., Cambridge, England, ν in cm-1).
1H and 13C-NMR spectra were recorded on a FT-NMR spec-
rometer (400 MHz) JNM-LA (Perkin Elmer Life and Analytical
Sciences, Inc., Boston, USA) using TMS as internal standard
(chemical shifts in ppm, δ units). Mass spectral data were
recorded on JEOL JMS-600H spectrometer (MA, USA). The
results of elemental analyses (C, H, N) were within 0.4% of
the theoretical values. Thin layer chromatography was per-
formed on silica gel GLF plates, 250 microns.
3-[4-(4-Aryl-3-cyano-1,2-dihydro-2-oxo-6-pyridyl)phenyl]-
sydnones (5aϪj)
Method 2: A mixture of 3-(4-acetylphenyl)sydnone (3) (0.61 g,
3 mmol), the appropriate aromatic aldehyde (3 mmol), ethyl
cyanoacetate (0.34 g, 3 mmol) and ammonium acetate
(1.85 g, 24 mmol) in absolute ethanol (30 ml) was heated
under reflux for 6 h. The reaction mixture was cooled to room
temperature. The separated solid was filtered, dried and
recrystallized from the suitable solvent.
Method 1: A mixture of 3-[4-(3-aryl-1-oxo-2-propen-1-yl)-
phenyl]sydnones 4aϪg (3 mmol), ethyl cyanoacetate (0.34 g,
3 mmol) and ammonium acetate (1.85 g, 24 mmol) in abso-
lute ethanol (30 ml) was heated under reflux for 6 h. The
reaction mixture was cooled to room temperature. The sepa-
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Arch. Pharm. Pharm. Med. Chem. 2004, 337, 164−170
Novel analogues of sydnone as potential antibacterial agents 169
Method 3: A mixture of compound 3 (0.61 g, 3 mmol), the
appropriate aromatic aldehyde (3 mmol), ethyl cyanoacetate
(0.34 g, 3 mmol) and ammonium acetate (1.85 g, 24 mmol)
was heated in an oil bath at 140Ϫ160°C for 30 min then
cooled to room temperature. Absolute ethanol was added
with stirring and the product was collected by filtration, dried
and recrystallized from the suitable solvent.
H, CH sydnone). 8g: 1.20 (s, 3H, CH3), 7.30Ϫ8.50 (m, 9H,
Ar-H, N=C-H, CH sydnone). 13C-NMR (DMSO-d6): 8c: 168.4
(CO, sydnone), 140.9 (CH3-C=N), 135.8 (N=CH), 135.3,
132.9, 129.8, 128.9, 128.3, 121.3 (Ar C), 94.8 (CH, sydnone).
MS, m/z (%), 8g: 375 (1.1, M+), 345 (5.2, M+-NO), 317 (63.8,
M+-CNO2), 290 (7.2, C15H12Cl2N2), 144 (26.5, C9H8N2),117
(46.1, C8H7N), 101 (100, C7H3N), 76 (97.5, C6H4).
3-[4-(2-Amino-4-aryl-3-cyano-6-pyridyl)phenyl]sydnones
(6aϪj)
2-Aroyl-1-[4-(ψ-5-oxo-1,2,3-oxadiazol-3-yl)phenyl]ethylidene-
hydrazine (9a-d) and Bis-[1-[4-(ψ-5-oxo-1,2,3-oxadiazol-3yl)-
phenyl]ethylidenehydrazinocarbonyl]methane (10)
Method 1: The same procedure as described in method 1 for
preparation of compound 5 was applied except that malono-
nitrile (0.20 g, 3 mmol) was used instead of ethyl cyano-
acetate. IR, 6a: 3439 (NH2), 3126 (CH, sydnone), 2209 (CN),
1748 (C=O, sydnone), 1637 (C=N). 6b: 3352 (NH2), 3150
(CH, sydnone), 2211 (CN), 1752 (C=O, sydnone), 1639 (C=
N). 6f: 3350 (NH2), 3149 (CH, sydnone), 2220 (CN), 1758
(C=O, sydnone), 1602 (C=N). 1H-NMR (DMSO-d6): 6a:
7.28Ϫ8.38 (m, 11H, Ar-H, CH sydnone), NH2 seemed to be
exchanged by the solvent. 6c: 6.70-7.90 (m, 10H, Ar-H, CH
sydnone). 6d: 6.77 (s, 1H, CH sydnone), 6.85-8.40 (m, 9H,
Ar-H). 6e: 3.81 (s, 3H, OCH3), 7.07-8.42 (m, 10H, Ar-H, CH
sydnone).
A mixture of 3-(4-acetylphenyl)sydnone (3) (2.04 g, 10 mmol),
and the appropriate acid hydrazide (10 mmol) in absolute
ethanol (30 ml) was heated under reflux for 8 h. The reaction
mixture was cooled to room temperature, the formed precipi-
tate was filtered, dried and recrystallized from the suitable
solvent. IR: 9a: 3311 (NH), 3120 (CH, sydnone), 1775 (C=O,
sydnone), 1662 (C=O, amide), 1588 (C=N). 9b: 3450 (OH),
3325 (NH), 3139 (CH, sydnone), 1729 (C=O, sydnone), 1676
(C=O, amide), 1602 (C=N). 9d: 3325 (NH), 3116 (CH, syd-
none), 1738 (C=O, sydnone), 1668 (C=O, amide), 1588 (C=
N). 10: 3334 (NH), 3100 (CH, sydnone), 1744 (C=O, syd-
none), 1683 (C=O, amide), 1600 (C=N). 1H-NMR (DMSO-d6):
9a: 1.19 (s, 3H, CH3), 6.69Ϫ8.31 (m, 10H, Ar-H, CH syd-
none), 10.13 (br s, 1H, NH; D2O exchangeable). 9c: 1.12 (s,
3H, CH3), 6.74Ϫ8.23 (m, 9H, Ar-H, CH sydnone), 10.79 (br
s, 1H, NH; D2O exchangeable). MS, m/z (%), 9a: 292 (5.7,
M+-NO), 264 (50.2, M+-CNO2), 237 (0.9, C15H13N2O), 131
(8.9, C8H7N2), 105 (100, C7H5O), 77 (82.9, C6H5).
Method 2: The same procedure as described in method 2 for
preparation of compound 5 was adopted except that malono-
nitrile (0.20 g, 3 mmol) was used instead of ethyl cyano-
acetate.
Method 3: The same procedure as described in method 3 for
preparation of compound 5 was followed except that malono-
nitrile (0.20 g, 3 mmol) was used instead of ethyl cyano-
acetate.
Antibacterial Activity
The synthesized compounds were evaluated for their in vitro
antibacterial activity against representative Gram-positive
and Gram-negative organisms applying standard agar di-
lution method using Mueller-Hinton medium. MICs were de-
termined after 18 h of incubation at 35°C. The concentrations
of the bacterial suspensions were verified by determining
standard colony counts on antibiotic-free agar plates. The
MIC was considered to be the lowest concentration that com-
pletely inhibited growth on agar plates disregarding a single
colony or a faint haze caused by the inoculum. Ciprofloxacin
was used as a reference compound.
1-[4-(ψ-5-Oxo-1,2,3-oxadiazol-3-yl)phenyl]ethylidene-
hydrazine (7)
Method 1: A mixture of compound 3 (2.04 g, 10 mmol), and
hydrazine hydrate 99% (5 g, 0.1 mol) in absolute ethanol (30
ml) was stirred for 8 h at room temperature. The product
formed was collected by filtration, dried and recrystallized
from ethanol-acetone mixture to yield 1.75 g (80%) of 7, mp
205Ϫ207°C. IR: 3381 (NH2), 3140 (CH, sydnone), 1750 (C=
O, sydnone), 1637 (C=N). 1H-NMR (DMSO-d6): 1.10 (s, 3H,
CH3), 4.92 (br s, 2H, NH2; D2O exchangeable), 6.84-8.44 (m,
5H, Ar-H, CH sydnone).
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Method 2: The same procedure as described in method 1
except that the reaction mixture was refluxed for 1 h. Further
work up as described in the previous method yielded 1.85 g
(85%) of 7, mp 206Ϫ208°C.
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