Efficient regiocontrolled synthesis and antimicrobial activity of pyrazoles

Article abstract of DOI:10.1055/s-2007-983771

A series of 1,5-diphenyl-1H-pyrazol-3-amines, 3-ethoxy-5-phenyl-1H- pyrazole, 5-ethoxy-1,3-diphenyl-1H-pyrazole and 3-ethoxy-1,5-diphenyl-1H- pyrazole were efficiently prepared from the regiocontrolled cyclization of α-oxoketene O,N-acetals and/or β-oxo thioxoesters with hydrazine derivatives using montmorillonite K-10 as solid support with ultrasound. The antimicrobial activity of the heterocyclic compounds was evaluated by direct bioautography test. Georg Thieme Verlag Stuttgart.

Full text of DOI:10.1055/s-2007-983771

PAPER
2485
Efficient Regiocontrolled Synthesis and Antimicrobial Activity of Pyrazoles
E
fficient
R
egiocontrol
a
led Synthe
s
r
is and A
a
ntimicrobial
A
ctiv
E
ity of Pyrazole
.
s
F. Braibante,* Hugo T. S. Braibante, Luciana de Carvalho Tavares, Simone F. Rohte, Carla C. Costa,
Ademir F. Morel, Caroline Z. Stüker, Robert A. Burrow
Departamento de Química, Universidade Federal de Santa Maria, 97105-900 Santa Maria, RS, Brazil
E-mail: mara@quimica.ufsm.br
Received 18 December 2006; revised 1 April 2007
H
H
H
N
Abstract: A series of 1,5-diphenyl-1H-pyrazol-3-amines, 3-
ethoxy-5-phenyl-1H-pyrazole, 5-ethoxy-1,3-diphenyl-1H-pyr-
N
N
N
N
Cl
R
azole and 3-ethoxy-1,5-diphenyl-1H-pyrazole were efficiently pre-
pared from the regiocontrolled cyclization of a-oxoketene O,N-
acetals and/or b-oxo thioxoesters with hydrazine derivatives using
montmorillonite K-10 as solid support with ultrasound. The antimi-
crobial activity of the heterocyclic compounds was evaluated by di-
rect bioautography test.
1
2a R = Me
2b R = Ph
2c R = Bn
O
2d R = CHMePh (R)
2e R = CHMePh (S)
2f R = H
2g R = allyl
Key words: pyrazoles, solid support K-10, regioselective synthe-
sis, bioautography
Figure 1 Structures of the heterocyclic compounds 1 and 2a–g
A series of 3-amino-substituted 1,5-diphenylpyrazoles
5a–e was synthesized regioselectively from the cycliza-
tion reaction of a-oxoketene O,N-acetals 4a–e with phe-
nylhydrazine on montmorillonite K-10, as solid support
under ultrasound (Scheme 1). The precursors 4a–e are po-
larized ketenes representing a new class of versatile func-
tionalized enamine carbonylic compound14 and were
prepared by reported procedures.13
Infections caused by microorganisms pose a serious chal-
lenge to the scientific community and the necessity for ef-
fective therapy has led to research in novel biologically
active agents. The pyrazole nucleus, among its numerous
pharmacological properties,1 also possesses antimicrobial
activity2 and, thus, is an ideal candidate. The synthesis of
1-arylpyrazoles usually involves the cyclocondensation of
1,3-dicarbonyl compounds and their equivalent 1,3-di-
electrophilic precursors such as b-dialkylamino-substitut-
ed chloro ketones with arylhydrazines. However, these
methods afford regioisomeric mixtures of pyrazoles.3,4 In
recent years, some success has been achieved in the regi-
oselective synthesis of 1-arylpyrazoles, however these
methods require severe reaction conditions5–8 and specific
bases.9
H
R
OH
S
O
N
RNH2
OEt
OEt
K-10
)))
4a–e
3
NH2NHPh
K-10
)))
a
b
c
d
e
R = Me
R = Ph
R = Bn
R = CHMePh (R)
R = CHMePh (S)
Our research group has been studying the synthesis of
polyfunctionalized heterocyclic systems utilizing reac-
tions on the solid support montmorillonite K-10.10–12 This
study presents the synthesis of a series of pyrazole com-
pounds using montmorillonite K-10 as a solid support un-
der solvent-free conditions. These heterocyclic systems
are similar to 5-(chloromethyl)-1H-pyrazole 1 and 5(3)-
H
N
N
N
R
5a–e
amino-substituted
3(5)-phenyl-1H-pyrazoles
2a–g
(Figure 1) which we have previously prepared.12,13
Scheme 1 Synthesis of the 3-amino-substituted 1,5-diphenylpyr-
azoles 5a–e
The heterocyclic 3-amino-substituted 1,5-diphenylpyra-
zoles 5a–e (Scheme 1), 3-ethoxy-5-phenyl-1H-pyrazole
(6) (Scheme 2) and 5-ethoxy-1,3-diphenyl-1H-pyrazole
(7) (Scheme 3) were synthesized and, together with com-
pounds 1 and 2a–g, their antibacterial and antifungal ac-
tivities were tested.
The structure of each product was identified from spectro-
scopic data. Due to the high selectivity observed in the
formation of the pyrazole rings 5a–e in heterogeneous
media, we decided to investigate the reactivity of the a-
oxoketene O,N-substituted acetal 4c (R = Bn) and unsub-
stituted 4f (R = H) with hydrazine and phenylhydrazine
hydrochloride using montmorillonite K-10 and ultrasound
sonication. The a-oxoketene O,N-acetals 4c,f were react-
ed with hydrazine hydrochloride under solid support and
ultrasound sonication to afford 3(5)-ethoxy-5(3)-phenyl-
SYNTHESIS 2007, No. 16, pp 2485–2490
x
x.
x
x
.
2
0
0
7
Advanced online publication: 12.07.2007
DOI: 10.1055/s-2007-983771; Art ID: M07306SS
© Georg Thieme Verlag Stuttgart · New York

2486
M. E. F. Braibante et al.
PAPER
1H-pyrazole 6 as shown in Scheme 2. The unexpected
loss of the amino group leading to the formation of hetero-
cyclic compound 6 is probably due to acidic conditions
from the presence of the hydrazine hydrochloride, making
this group a better leaving group that the ethoxy group.
When the same starting materials 4c and 4f were used in
the reaction with hydrazine hydrate in its free form, the
aminopyrazoles 2c,g were produced.
N
O
S
N
NH2NHPh⋅HCl
O
OEt
K-10, 24 h
)))
3
7
OH
S
H
R
N
O
N
N
NH2NH2⋅HCl
OEt
NH2NHPh⋅HCl
O
H
K-10, 20 h
)))
OEt
K-10, 19 h
)))
3
N
N
8
4c R = Bn
4f R = H
O
Scheme 3 Synthesis of the 5-ethoxy-1,3-diphenyl-1H-pyrazole (7)
and 3-ethoxy-1,5-diphenyl-1H-pyrazole (8)
6
H
R
NH2NH2⋅HCl
O
N
K-10, 22 h
)))
pounds 5a and 5c and Figure 3 shows the two crystallo-
graphically independent molecules, labeled I and II, found
in the asymmetric unit of compound 7. All compounds
contain a central pyrazole ring with three substituents, an
N-phenyl group, a C-phenyl group and a secondary amine
(5a and 5c) or ethyl ether moiety (7). The X-ray study
confirms that in compounds 5a and 5c, the C-phenyl
group is in the 5-position, adjacent to the N-phenyl group,
and the secondary amine is in the 3-position, while in
compound 7, the C-phenyl is in the 3-position and the eth-
yl ether is in the 5-position. The regiochemistry differenc-
es support the different mechanism of attack and
cyclization. In the structures, the pyrazole ring system
shows a delocalized double bond system that is almost ex-
actly planar (root-mean-square deviations less than 0.005
Å). An analysis of the bond lengths within the pyrazole
ring show that they are consistent with average bonds
lengths found in the literature for N-phenylpyrazole com-
pounds (42 observations) obtained from the Cambridge
Structural Database16 (Table 1).17
OEt
4c,j
Scheme 2 Synthesis of the 3(5)-ethoxy-5(3)-phenyl-1H-pyrazole 6
Due to loss of the amino group in the formation of the
pyrazole 6, we decided to repeat the cyclization reaction
using the a-oxo thioxoester 3, instead, as starting material
to evaluate its reactivity with the reaction conditions used
(Scheme 2). The same 1H-pyrazole 6 was obtained. We
believed that the keto form of the compound 3 may be
present in the reaction conditions used, so that through
this tautomeric form the formation of pyrazole 6 is possi-
ble. The structure of the compound 6 was identified from
spectroscopic data.
Continuing the study of the reactivity of the a-oxo thioxo-
ester 3 and of the O,N-acetals 4c,f, we performed the re-
action of these compounds with phenylhydrazine
hydrochloride using solid support and ultrasound to gen-
erate selectively the regioisomers 5-ethoxy-1,3-diphenyl-
1H-pyrazole (7) and 3-ethoxy-1,5-diphenyl-1H-pyrazole
(8) (Scheme 3). Compound 7 is obtained by probable ini-
tial attack of the more nucleophilic nitrogen of phenylhy-
drazine hydrochloride at the carbonylic carbon of b-oxo
thioxoester 3, with loss of water followed by cyclization.
The formation of compound 8 is considered to proceed by
initial attack of the more nucleophilic nitrogen of phenyl-
hydrazine hydrochloride at Cb of O,N-acetals 4c,f, with
loss of the amino group followed by intramolecular cy-
clization.
The antimicrobial activity of 5-(chloromethyl)pyrazole 1,
3-amino-substituted 5-phenyl-1H-pyrazoles 2a–g, 3-ami-
no-substituted 1,5-diphenyl-1H-pyrazoles 5a–e, 3-eth-
oxy-5-phenyl-1H-pyrazole (6) and 5-ethoxy-1,3-diphen-
yl-1H-pyrazole (7) were evaluated by direct
bioautography18 using a TLC bioassay against microor-
ganisms strains. The collection of eleven microorganisms
used included three Gram-positive bacteria: Staphylococ-
cus aureus (Sa), Staphylococcus epidermidis (Se), Bacil-
lus subtilis (Bs); four Gram-negative bacteria:
Eschericchia coli (Ec), Salmonella setubal (Ss),
Pseudomonas aeruginosa (Pa), Klebsiella pneumoniae
(Kp); four fungi: Candida albicans (Ca), Saccharomyces
cerevisiae (Sc), Cryptococcus neoformans (Cn), and Can-
dida dubliniensis (Cd- isolated clinical). For the antimi-
crobial analysis, initially we compared compounds 1 and
2d, which possess different patterns of substitution.
Among the series of the aminopyrazoles 2a–g, we chose
the heterocycle 2d, that has an asymmetrical center [(R)-
CHMePh].
The structures of the regioisomers 7 and 8 were unambig-
uously established by 1H and 13C NMR spectra. The pre-
viously reported synthesis15 of regioisomeric compounds
similar to 7 and 8 requires specific reaction conditions
such as elevated temperature and basic media.
X-ray crystallographic studies on compounds 5a, 5c, and
7 were undertaken to confirm the regiochemistry of the
products. Figure 2 shows the molecular structures of com-
Synthesis 2007, No. 16, 2485–2490 © Thieme Stuttgart · New York

PAPER
Efficient Regiocontrolled Synthesis and Antimicrobial Activity of Pyrazoles
2487
Figure 3 X-ray crystal structure of the two crystallographically
independent molecules of 5-ethoxy-1,3-diphenyl-1H-pyrazole (7)
Based on the promising bioactive behavior presented by
the series of the aminopyrazoles 2a–g, we decided to in-
vestigate the structure–activity relationship of their 3-
amino-substituted 1,5-diphenylpyrazoles 5a–e analogues,
as well as of the heterocycles 3-ethoxy-5-phenyl-1H-
pyrazole (6) (Scheme 2) and 5-ethoxy-1,3-diphenyl-1H-
pyrazole (7) (Scheme 3). The bioassay of compounds 5a–
e shows that these heterocycles are inactive against all
tested microorganisms. This finding clearly indicates that
the presence of the phenyl substituent in the N1 position
of the ring is responsible for the inactivity of these com-
pounds. An identical result to those described above was
observed with the ethoxypyrazole 7, which also shows no
antimicrobial activity against all the analyzed microor-
ganisms.
Figure 2 X-ray crystal structure of 3-(methylamino)-1,5-diphenyl-
1H-pyrazole (5a) and 3-(benzylamino)-1,5-diphenyl-1H-pyrazole
(5c)
The bioassay of compound 1 shows that it presents con-
centration dependent activity against all tested microor-
ganisms. Good antifungal activity is observed against all
analyzed fungi, with strong inhibition against C. dublin-
iensis (1.0 mg) compared to the control. Compound 2d ex-
hibits promising antibacterial activity against all Gram-
positive and Gram-negative bacteria (Table 2). It is possi-
ble to detect the same antibacterial tendency shown by the
heterocycle 2d in the remaining compounds of the series
of the 5-amino-substituted pyrazoles [R = Me, Ph, Bn,
(S)-CHMePh, H, allyl] (Table 1). In particular, compound
2e exhibits very good antibacterial potential (0.15 mg)
against all strains of tested bacteria, however, it is inactive
against all the analyzed fungi in the same way as com-
pound 2a. The excellent results of the bacterial inhibition
demonstrated by 2e compared that of its isomer 2d, and
with the other analyzed aminopyrazoles, suggests that the
stereochemistry [(S)-CHMePh] present in structure of 2e
plays an important role.
The phenyl group in the N1 position continues to inhibit
biological activity despite the different substituents on the
pyrazole ring. On the other hand, the heterocyclic com-
pound 6 demonstrates moderate antibacterial activity
against the tested bacteria (Table 3), when compared to
the group control and the series 2a–g, except for the bac-
terium Gram-negative K. pneumoniae for which was not
active, but, similar to 2a, 2e, and 5a–e, it is inactive
against fungi. The fact that ethoxypyrazole 6 presents bac-
terial inhibition against all Gram-positive bacteria and
Synthesis 2007, No. 16, 2485–2490 © Thieme Stuttgart · New York

2488
M. E. F. Braibante et al.
PAPER
Table 1 Bond Distances within the Pyrazole Ring for 5a, 5c, and 7a
Compound
Bond lengths (Å)
N1–N2
N1–C5
N2=C3
C3–C4
C4=C5
5a
1.3887(14)
1.3787(13)
1.3887(14)
1.3766(15)
1.371(13)
1.366(19)
1.3698(16)
1.3596(14)
1.3666(19)
1.3628(20)
1.363(12)
1.336(17)
1.3337(15)
1.3282(14)
1.3243(20)
1.3255(19)
1.330(9)
1.4075(19)
1.4061(15)
1.4043(19)
1.4064(21)
1.400(14)
1.513(14)
1.3634(19)
1.3726(16)
1.3595(19)
1.3651(19)
1.366(11)
1.299(27
5c
7(I)
7(II)
average, N-phenylpyrazoles
average, all compounds17
1.279(8)
a With comparisons to the averages found for N-phenylpyrazole rings (42 observations in the C.S.D.) and for N–N, N–C, N=C, C–C, and C=C
bonds for all compounds.
Table 2 Bioautography Assay Results (mg) of 1 and 2a–g
Microorganisms
1
2a
2b
2c
2d
2e
2f
2g
0.5
Controla
0.7
Staphylococcus aureus
ATCC 6538p
50
0.75
3.12
1.56
3.12
1.0
3.12
1.0
na
0.75
0.15
0.5
0.5
6.25
1.0
0.5
1.0
0.15
0.5
3.12
na
0.15
0.15
0.15
0.15
0.15
0.15
0.15
na
3.12
3.12
6.25
3.12
3.12
6.25
6.25
na
Staphylococcus epidermidis ATCC 12228
100
25
0.75
3.12
3.12
1.0
25
0.7
Bacillus subtilis
ATCC 6633
ATCC 25792
ATCC 19796
ATCC 10031
ATCC 27853
ATCC 2601
ATCC 10231
ATCC 28952
12.5
0.5
12.5
12.5
12.5
na
0.8
Eschericchia coli
100
50
0.15
0.15
0.15
0.75
na
0.5
Salmonella setubal
0.7
Klebsiella pneumoniae
Pseudomonas aeruginosa
Saccharomyces cerevisiae
Candida albicans
50
0.15
1.56
na
0.5
100
6.25
6.25
3.12
0.6
3.24
2.43
2.43
4.05
na
na
na
na
na
na
100
na
Cryptococcus neoformans
Candida dubliniensis
na
0.75
25
25
na
na
100
na
isolated clinical SM-26 1.0
na
6.25
25
na
na
a Standard antibiotic chloramphenicol (bacteria) and Nistatine (fungi) (mg); na = not active.
against three Gram-negative bacteria, is due to the lack of diphenylpyrazoles 5a–e and 5-ethoxy-1,3-diphenyl-1H-
the phenyl substituent in the position N1 of the pyrazole pyrazole (7) shows that the presence of the amino group
ring, similar the aminopyrazoles 2a–g, which are active in the ring enhances the antibacterial activity against the
against all tested bacteria.
tested bacteria while the presence of the phenyl in the N1
position substituent on the pyrazole ring inhibits activity
against all of the employed microorganisms.
In summary, the reactions using montmorillonite K-10
support and ultrasound provides a highly selective method
for the synthesis of 1,3- and 1,5-diphenylpyrazoles. The
results of the in vitro biological evaluation against various
microorganisms indicates that 5-(chloromethyl)pyrazole
1 demonstrates good antifungal behavior and the series
5(3)-amino-substituted 3(5)-phenyl-1H-pyrazoles 2a–g
presents strong antibacterial behavior. Furthermore, the
structure–activity-relationship study comparing the re-
sults of the bioactive aminopyrazoles 2a–g, and ethoxy-
Melting points were determined with a Microquímica APF-301 ap-
paratus and are uncorrected. 1H and 13C NMR spectra were recorded
on a Bruker DPX 200 and 400 spectrometer in CDCl3/TMS. Crys-
tallographic measurements were made on a Bruker X8 Kappa Apex
II CCD area detector equipped automatic diffractometer with
MoKa radiation (l = 0.71073 Å). Structures were solved by direct
methods (SHELXS-97) and additional atoms were located in the
difference Fourier map and refined on F2 (SHELXL-97). An ultra-
pyrazole 6, with the bioinactive series 3-amino-1,5- sound bath (water), Thornton 50–60 Hz, 110/220 V, 1.0 amps was
used. The temperature of the water bath never exceeded 35 °C.
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PAPER
Efficient Regiocontrolled Synthesis and Antimicrobial Activity of Pyrazoles
2489
1,5-Diphenyl-3-{[(R)-1-phenylethyl]amino}-1H-pyrazole (5d)
and 1,5-Diphenyl-3-{[(S)-1-phenylethyl]amino}-1,5-diphenyl-
1H-pyrazole (5e)
Compound 5d
Chromatography: 10% EtOAc–hexane; oil; yield: 10%.
Table 3 Bioautography Assay Results (mg) of 6
Microorganisms
6
Controla
0.7
Staphylococcus aureus
ATCC 6538p
50
Staphylococcus epidermidis ATCC 12228
3.12
12.5
12.5
50
0.7
Compound 5e
Chromatography: 25% EtOAc–hexane; oil; yield: 21%.
Bacillus subtilis
ATCC 6633
ATCC 25792
ATCC 19796
ATCC 10031
ATCC 27853
ATCC 2601
ATCC 10231
ATCC 28952
0.8
Compounds 5d and 5e
Eschericchia coli
0.5
1H NMR (400 MHz, CDCl3): d = 7.05–7.33 (m, 15 H), 5.55 (s, 1 H),
4. 58 (br, 1 H), 4.46 (q, J = 6.8 Hz, 1 H), 1.45 (d, J = 6.8 Hz, 3 H).
Salmonella setubal
0.7
13C NMR (100 MHz, CDCl3): d = 156.37, 145.21, 143.74, 139.80,
130.62, 128.62, 128.54, 128.45, 128.21, 128.10, 126.87, 126.36,
126.04, 124.56, 94.38, 54.57, 24.35.
Klebsiella pneumoniae
Pseudomonas aeruginosa
Saccharomyces cerevisiae
Candida albicans
na
0.5
25
0.6
na
3.24
2.43
2.43
4.05
3-Ethoxy-5-phenyl-1H-pyrazole (6)
PhNHNH2·HCl (10 mmol) was added to the a-oxoketene O,N-ace-
tal 4c or 4f or to the a-oxo thioxoester 3 (5 mmol) dispersed on
montmorillonite K-10, the mixture was placed in ultrasound bath
for 20 h or 22 h. The product was extracted by washing the mont-
morillonite K-10 with CH2Cl2; the CH2Cl2 soln was dried (MgSO4),
filtered, and the solvent was removed in vacuo to yield the crude
product. The compound was purified by column chromatography
(silica gel, Aldrich 70–230 mesh, 15% EtOAc–petroleum ether) to
give analytically pure pyrazole as a solid; yield: 43% (from 4c),
59% (from 4f), and 47% (from 3); mp 127–129 °C.
na
Cryptococcus neoformans
Candida dubliniensis
na
isolated clinical
SM-26
na
a Standard antibiotic chloramphenicol (bacteria) and Nistatine (fungi)
(mg); na = not active.
1H NMR (400 MHz, CDCl3): d = 7.32–7.60 (m, 5 H), 5.93 (s, 1 H),
4.22 (q, J = 6.8 Hz, 2 H), 1.37 (t, J = 6.8 Hz, 3 H).
13C NMR (100 MHz, CDCl3): d = 163.96, 144.95, 129.98, 128.87,
128.44, 125.36, 87.93, 65.00, 14.84.
3-Amino-Substituted 1,5-Diphenylpyrazoles 5a–e; General
Procedure
Phenylhydrazine (10 mmol) was added to the appropriate a-oxo-
ketene O,N-acetals 4a–e (5 mmol) dispersed on montmorillonite
K-10, the mixture was placed in ultrasound bath for 19 h. The prod-
uct was extracted by washing the montmorillonite K-10 with
CH2Cl2; the CH2Cl2 soln was dried (MgSO4), filtered, and the sol-
vent was removed in vacuo to yield the crude product. All the com-
pounds were purified by column chromatography (silica gel,
Aldrich 70–230 mesh, EtOAc–hexane) to give analytically pure
pyrazoles.
5-Ethoxy-1,3-diphenyl-1H-pyrazole (7)
PhNHNH2·HCl (10 mmol) was added to the a-oxo thioxoester 3 (5
mmol) dispersed on montmorillonite K-10 and the mixture was
placed in ultrasound bath for 24 h. The product was extracted by
washing the montmorillonite K-10 with CH2Cl2; the CH2Cl2 soln
was dried (MgSO4), filtered, and the solvent was removed in vacuo
to yield the crude product. The compound was purified by column
chromatography (silica gel, Aldrich 70–230 mesh, 15% EtOAc–
hexane) to give analytically pure 7 as a solid; yield: 52%; mp 67–69
°C.
1H NMR (400 MHz, CDCl3): d = 7.23–7.86 (m, 10 H), 5.97 (s, 1 H),
4.21 (q, J = 7.0, 2 H), 1.46 (t, J = 7.0, 3 H).
13C NMR (100 MHz, CDCl3): d = 155.24, 150.43, 138.77, 133.32,
128.73, 128.47, 127.97, 126.04, 125.49, 122.02, 83.74, 68.02,
14.56.
3-(Methylamino)-1,5-diphenyl-1H-pyrazole (5a)
Chromatography: 30% EtOAc–hexane; solid; yield: 39%; mp 107–
109 °C.
1H NMR (200 MHz, CDCl3): d = 7.19–7.28 (m, 10 H), 5.92 (s, 1 H),
3.94 (br, 1 H), 2.93 (s, 3 H).
13C NMR (100 MHz, CDCl3): d = 157.62, 144.14, 139.96, 130.69,
128.72, 128.58, 128.31, 128.18, 126.48, 124.76, 94.56, 31.57.
1,5-Diphenyl-3-(phenylamino)-1H-pyrazole (5b)
Chromatography: 25% EtOAc–hexane; oil; yield: 43%.
3-Ethoxy-1,5-diphenyl-1H-pyrazole (8)
PhNHNH2·HCl (10 mmol) was added to the a-oxoketene O,N-ace-
tal 4c or 4f (5 mmol) dispersed on montmorillonite K-10, the mix-
ture was placed in ultrasound bath for 19 h. The product was
extracted by washing the montmorillonite K-10 with CH2Cl2; the
CH2Cl2 soln was dried (MgSO4), filtered, and the solvent was re-
moved in vacuo to yield the crude product. The compound was pu-
rified by column chromatography (silica gel, Aldrich 70–230 mesh,
10% EtOAc–hexane) to give analytically pure 8 as an oil; yield:
38% (from 4c) and 63% (from 4f).
1H NMR (200 MHz, CDCl3): d = 7.19–7.31 (m, 10 H), 5.95 (s, 1 H),
4.31 (q, J = 7.0 Hz, 2 H), 1.43 (t, J = 7.0 Hz, 3 H).
13C NMR (100 MHz, CDCl3): d = 163.53, 144.11, 140.11, 130.77,
128.74, 128.69, 128.37, 128.27, 126.61, 124.93, 93.69, 64.71,
14.88.
1H NMR (400 MHz, CDCl3): d = 7.19–7.32 (m, 15 H), 6.31 (br, 1
H), 6.26 (s, 1 H).
13C NMR (100 MHz, CDCl3): d = 151.97, 143.60, 142.81, 139.97,
130.59, 130.59, 129.17, 128.74, 128.68, 128.40, 128.30, 126.68,
124.77, 120.10, 116.13, 97.23.
3-(Benzylamino)-1,5-diphenyl-1H-pyrazole (5c)
Chromatography: 10% EtOAc–hexane; solid; yield: 19%; mp 114–
116 °C.
1H NMR (400 MHz, CDCl3): d = 7.14–7.42 (m, 15 H), 5.83 (s, 1 H),
4.42 (s, 2 H), 3.87 (br, 1 H).
13C NMR (100 MHz, CDCl3): d = 157.19, 143.84, 140.14, 139.81,
130.85, 128.61, 128.56, 128.42, 128.25, 128.05, 127.54, 127.04,
126.25, 124.62, 94.73, 48.96.
Synthesis 2007, No. 16, 2485–2490 © Thieme Stuttgart · New York

2490
M. E. F. Braibante et al.
PAPER
Crystallographic Measurements19
Hydrogen atoms were found in the difference Fourier map but re-
Acknowledgment
We are grateful to the Conselho Nacional de Desenvolvimento
Científico e Tecnológico (CNPq -PIBIC), Brazil for fellowships to
S.F.R.
fined in idealized geometric positions (Cphenyl–H = 0.93; Cmethylene
–
H = 0.97; N–H = 0.89 Å). The space groups were determined by
systematic absences and overall intensity statistics for compounds
5a and 5c. For compound 7, the space group could not be easily de-
termined. Crystalline metric and Laue symmetry suggested a mon-
oclinic C space group without any additional translational
symmetry. Intensity statistics suggested a noncentrosymmetric
space group. Direct methods solutions (SHELXS-97) were tried un-
successfully in C2, Cm and C2/m without any satisfactory result; the
space groups possessed inappropriate additional symmetry causing
the found molecules to fuse together. An acceptable soln was found
in the primitive space group, P1, at half the volume of the C cen-
tered one. ROTAX20 found a twofold axis about [–1 1 0]. Applying
the twin law [0 –1 0 –1 0 0 0 0 –1] allowed the satisfactory comple-
tion of the refinement of the structure.
References
(1) (a) Manfredini, S.; Bazzanini, R.; Baraldi, P. G.; Guarneri,
M.; Simoni, D.; Marongiu, M. E.; Pani, A.; Tramontano, E.;
Colla, P. L. J. Med. Chem. 1992, 35, 917. (b) Kees, K. L.;
Fitzgerald, J. J.; Steiner, K. E.; Mattes, J. F.; Mithan, B.;
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Crystal data for 5a: C16H15N3, M = 249.31, orthorhombic, space
group Pbca (No. 61), a = 8.5449(5) Å, b = 12.2609(9) Å,
c = 26.0612(17) Å, V = 2730.4(3) Å3, T = 294(2) K, Z = 8,
(2) (a) Pinemova, E. V.; Voronina, É. V. Pharm. Chem. J. (Engl.
Transl.) 2001, 35, 602. (b) Kane, J. L.; Hirth, B. H.; Liang,
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Dc = 1.213 g/cm–3, m = 0.074 mm–1, 3.01
< q < 29.61°,
F(000) = 1056; 15574 reflections measured, 3785 unique
(Rint = 0.0521). The final wR2 = 0.1175 (all data), R1 [I >
s(I)] = 0.0429, GoF = 0.905. CCDC No. 611960.
Crystal data for 5c: C22H19N3, M = 325.40, monoclinic, space group
P21/c (No. 14), a = 5.6704(2) Å, b = 19.0708(8) Å, c = 16.5995(7)
Å, b = 98.942(2)°, V = 1773.24(12) Å3, T = 294(2) K, Z = 4,
Dc = 1.219 g/cm–3, m = 0.073 mm–1, 3.44
< q < 29.74°,
(4) Kashima, C.; Harada, H.; Kita, I.; Fukuchi, I.; Hosomi, A.
F(000) = 688; 20995 reflections measured, 5020 unique
(Rint = 0.0299). The final wR2 = 0.1406 (all data), R1 [I >
s(I)] = 0.0444, GoF = 1.013. CCDC No. 611961.
Synthesis 1994, 61.
(5) Sakya, S. M.; Rast, B. Tetrahedron Lett. 2003, 44, 7629.
(6) Moreno-Manãs, M.; Sebastián, R. M.; Vallribera, A.; Carini,
F. Synthesis 1999, 157.
(7) Lee, K. Y.; Kim, J. M.; Kim, J. N. Tetrahedron Lett. 2003,
44, 6737.
(8) Huang, Y. R.; Katzenellenbogen, J. A. Org. Lett. 2000, 2,
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(9) Peruncheralathan, S.; Yadav, A. K.; Ila, H.; Junjappa, H.
J. Org. Chem. 2005, 70, 9644.
Crystal data for 7: C17H16N2O, M = 264.32, triclinic, space group
P1 (No. 2), a = 9.7703(2) Å, b = 9.7746(2) Å, c = 15.3970(3) Å,
a = 97.0210(10)°,
b = 97.0020(10)°,
g = 93.0100(10),
V = 1445.06(5) Å3, T = 294(2) K, Z = 4, Dc = 1.215 g/cm–3,
m = 0.077 mm–1, 1.34 < q < 30.07°, F(000) = 560; 63439 reflections
measured, 8442 unique (Rint = 0.0291). The final wR2 = 0.1314 (all
data), R1 [I > s(I)] = 0.0425, GoF = 1.042. CCDC No. 611962.
(10) Braibante, M. E. F.; Braibante, H. T. S.; Valduga, C. J.
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Antimicrobial Activities
The antimicrobial activities of the compounds 1, 2a–g, 3a–e, 6, and
7 were assayed using the bioautography technique.18 The collection
of eleven microorganisms used included three Gram-positive bacte-
ria: Staphylococcus aureus (ATCC 6538p), Staphylococcus epider-
midis (ATCC 12228), and Bacillus subtilis (ATCC 6633); four
Gram-negative bacteria: Eschericchia coli (ATCC 25792), Salmo-
nella setubal (ATCC 19796), Pseudomonas aeruginosa (ATCC
27853), and Klebsiella pneumoniae (ATCC 10031); four fungi:
Candida albicans (ATCC 10231), Saccharomyces cerevisiae
(ATCC 2601), Cryptococcus neoformans (ATCC 28952), and Can-
dida dubliniensis (isolated clinical SM-26). Standard microorgan-
ism strains were maintained at the Chemistry Department of the
University of Santa Maria, RS, Brazil. For bioassay, the compounds
were applied to pre-coated TLC plates in concentrations from 100
to 0.15 mg. Mueller–Hinton agar medium (MHA-Merck) was inoc-
ulated with microorganisms suspended in saline soln (105 CFU/mL)
and distributed over TLC plates. Bacterium and yeast plates were
incubated at 37 °C for 24 h and at 25 °C for 72 h, respectively. Stan-
dard antibiotic chloramphenicol and nistatine were used to control
the sensitivity of the microbial test. After incubation, the plates
were stained with an aqueous soln of 2,3,5-triphenyltetrazolium
chloride (TTC, 1 mg/mL). The appearance of inhibition zones was
used to demonstrate the lesser sample amount that inhibited micro-
organism growth. Samples were tested in triplicate.
(11) Braibante, M. E. F.; Braibante, H. T. S.; Valduga, C. J.;
Santis, D. B. J. Heterocycl. Chem. 1999, 36, 505.
(12) Braibante, M. E. F.; Braibante, H. T. S.; Costa, C. C.;
Martins, D. B. Tetrahedron Lett. 2002, 43, 8079.
(13) Braibante, M. E. F.; Braibante, H. T. S.; Roza, J. K.;
Henriques, D. M.; Tavares, L. C. Synthesis 2003, 1160.
(14) Mahata, P. K.; Venkatesh, C.; Kumar, U. K. S.; Ila, H.;
Junjappa, H. J. Org. Chem. 2003, 68, 3966.
(15) Peruncheralathan, S.; Khan, T. A.; Ila, H.; Junjappa, H.
J. Org. Chem. 2005, 70, 10030.
(16) Allen, F. H. Acta Crystallogr., Sect. B 2002, 58, 380.
(17) Allen, F. H.; Kennard, O.; Watson, D. G.; Brammer, L.;
Orpen, A. G. J. Chem. Soc., Dalton Trans. 1987, S1.
(18) Rahalison, L.; Hamburger, M.; Hostettman, K.; Monod, M.;
Frank, E. Phytochem. Anal. 1991, 2, 199.
(19) The CCDC deposits contain the supplementary
crystallographic data for this paper. These data can be
obtained free of charge from The Cambridge
Crystallographic Data Centre via www.ccdc.cam.ac.uk/
data_request/cif.
(20) ROTAX, Version 26th November 2001; Parsons, S.; Gould,
R.; University of Edinburgh: UK, with additions by Cooper,
R. (Oxford) and Farrugia, L. (Glasgow).
Synthesis 2007, No. 16, 2485–2490 © Thieme Stuttgart · New York