A convenient synthesis, reactions and biological studies of some novel selenolo[2,3-c]pyrazole compounds as antimicrobial and anti-inflammatory agents

Article abstract of DOI:10.1007/s00044-016-1536-8

5-Chloro-3-methyl-1-phenylpyrazole-4-carbonitrile 3 was reacted with selenium in the presence of sodium borohydride and chloroacetamide to afford selanyl acetamide 5, which underwent Thorpe-Ziegler cyclization upon heating with sodium ethoxide to give the

Full text of DOI:10.1007/s00044-016-1536-8

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-016-1536-8
ORIGINAL RESEARCH
A convenient synthesis, reactions and biological studies of some
novel selenolo[2,3-c]pyrazole compounds as antimicrobial
and anti-inflammatory agents
1
1
1
Remon M. Zaki
•
Yasser A. El-Ossaily
•
Ahmed A. Geies
Received: 24 October 2015 / Accepted: 11 February 2016
Ó Springer Science+Business Media New York 2016
Abstract 5-Chloro-3-methyl-1-phenylpyrazole-4-carboni-
trile 3 was reacted with selenium in the presence of sodium
borohydride and chloroacetamide to afford selanyl acetamide
showed remarkable antibacterial, antifungal and anti-in-
flammatory activities.
CN
NH
2
5
, which underwent Thorpe–Ziegler cyclization upon heating
with sodium ethoxide to give the novel synthesized 4-amino-
-methyl-1-phenyl-1H-selenolo[2,3-c]pyrazole-5-carbox-
NH
2
3
N
N
Cl
N
amide compound (6). The latter compound was used as a
versatile precursor for synthesis of other heterocyclic rings,
namely pyrimidine, imidazopyrimidine and thiadi-
azinopyrimidine fused to selenolo[2,3-c]pyrazole moiety.
The newly synthesized compounds and their derivatives
were characterized by elemental and spectral analysis (IR,
N
Se
O
1
13
H NMR, C NMR and mass spectrometric analyses).
Furthermore, some of these synthesized compounds were
screened against various pathogenic bacterial and fungal
strains. The results demonstrate that most of the synthe-
sized compounds possess a significant antibacterial activity
against gram-positive and gram-negative bacteria. Also,
some of these compounds showed a remarkable antifungal
activity, especially Candida albicans. On the other hand,
some of the synthesized compounds possess high anti-in-
flammatory activity using carrageenan-induced rat paw
edema assay compared with indomethacin.
N
N
R
N
N
N
Se
O
Graphical Abstract The present work discussed synthesis
of new selenolo[2,3-c]pyrazoles fused to other heterocyclic
rings, namely pyrimidine, imidazopyrimidine and thiadi-
azinopyrimidine. Some of the synthesized compounds
Keywords Synthesis Á Reactions Á Selenolopyrazole Á
Pyrimidine Á Antimicrobial activity Á
Anti-inflammatory activity
Introduction
&
Remon M. Zaki
remonch2003@yahoo.com;
rasal@aun.edu.eg
Pyrazoles/N-arylpyrazoles are very interesting class of
heterocyclic compounds which were considered as scaf-
folds of many efficacious biologically active agents such as
antitumor (Barnes et al., 2001; Altintop et al., 2014; Bar-
aldi et al., 2002; Mercep et al., 2004), anti-inflammatory
1
Chemistry Department, Faculty of Science, Assiut
University, Assiut 71516, Egypt
123

Med Chem Res
(
Smith et al., 2001; Raffa et al., 2010; Maggio et al., 2001;
Cardia et al., 1998; Mullican et al., 1993; Ochi et al.,
001), hypoglycemic (Maekawa et al., 2004), antithrom-
botic (Qiao et al., 2004), adenosine receptor antagonist
Baraldi et al., 2003), kinase inhibitory activity (Brown
wafer technique) with a Perkin-Elmer 1430 Spectropho-
1
13
tometer. H NMR and CNMR spectra were obtained on a
Varian EM-390 MHz (90 MHz) and Bruker 400 MHz
spectrometers in CDCl , DMSO-d and CF CO D using
2
3
6
3
2
(
Me Si as internal standard, and chemical shifts are
4
et al., 2004), insecticidal (Stevensons et al., 2004),
ulcerogenic (Ochi et al., 2001), neuropeptide Y receptor
type 5 (Stamford and Wu, 2004) and thrombopoietin
mimetics activities (Heerding 2004). Members of this class
of compounds have also been investigated for their local
anesthetic, antiarrhythmic (Iovu et al., 2000), herbicidal
expressed as ppm. Mass spectrometric analysis was mea-
sured on a Joel-JMS 600 mass spectrometer. Analytical
data were obtained on elemental analyzer system GmbH-
VarioEL V.3 microanalyzer in the central laboratory of
Assiut University, and all results were found to be in an
acceptable range (± 0.20). The substituted pyrazole com-
pounds 1–3 were synthesized according to the literature
procedure (Haider et al., 2005), with m.p. 140–142 °C for
compound 1. Some of the synthesized compounds in this
work were chosen and screened in vitro for their antimi-
crobial activity against some strains of bacteria and fungi.
The antifungal and antibacterial activities of tested com-
pounds were evaluated by the reported method (Kwon-
Chung and Bennett, 1992) using 0.005 % (100 lg/2 mL)
concentration of selected compounds in DMSO as a sol-
vent. The inhibition zone (mm) was compared with levo-
floxacin and clotrimazole as a reference for antibacterial
and antifungal activities, respectively. The results of anti-
inflammatory effect of some synthesized compounds were
analyzed by one-way analysis of variance (ANOVA) fol-
lowed by Newman–Keuls multiple comparison test as a
post-test. These analyses were carried out using computer
Prism program for Windows version 3.0 (GraphPad soft-
ware, Inc, San Diego CA, USA). The significant difference
between groups was accepted at P \ 0.05 *, 0.01** or
0.001***, and the data were expressed as mean ± standard
error (SE).
(
Vicentini et al., 1999), antiviral (Storer et al., 1999; El-
Sabbagh et al., 2009), immunosuppressant (Wang et al.,
998), anticonvulsant (El-Sabbagh et al., 2009), antide-
1
pressant (Abdel-Aziz et al., 2009) and antimicrobial
activities (Lee et al., 2001; Sridhar et al., 2004; Abdel-
Gawad et al., 2003; Mamolo et al., 2001; Soliman et al.,
2
001; Mishriky et al., 2001). On the other hand, selenium
has attracted the attention of many scientists working in a
variety of fields due to its dietary antioxidant activity and is
an essential component of active sites of several enzymes
(
Richardson et al., 2006). Literature survey revealed that
the most representative biological activity among fused
pyrazoles with five-membered heterocycles such as
selenolopyrazole is the antitumor activity (Raffa et al.,
2
015; Chou et al., 2010).
In the present work, we synthesized new heterocyclic
rings such as imidazopyrimidine and thiazdiazinopyrim-
idine fused to selenolopyrazole moiety. Imidazopyrimidi-
nes are important scaffolds in medicinal chemistry, as they
exhibit a broad spectrum of biological activity such as
anticancer (Cosimelli et al., 2014), antitubercular (Mar-
giotta et al., 2007), antiviral (Gueiffier et al., 1996),
antimicrobial (Rival et al., 1992), antifungal (Rival et al.,
5
-Chloro-3-methyl-1-phenyl-1H-pyrazole-4-carbaldehy-
doxime (2) A mixture of the chloro-aldehyde 1 (11 g,
1
991), anti-inflammatory (Vidal et al., 2001), parasiticidal
activity (Jiricek et al., 2014), calcium channel blockers
Sanfilippo et al., 1988) and benzodiazepine receptor
0
0
.05 mol) and hydroxylamine hydrochloride (4.20 g,
.06 mol) in ethanol (100 mL) in the presence of fused
(
sodium acetate (5 g, 0.06 mol) as basic catalyst, was
refluxed for 2 h. The solid product, which was formed on
cooling and poured into ice-water mixture, was recrystal-
lized from ethanol as white crystals (93.40 %, 11 g) yield;
mp 130–132 °C; IR (KBr) mmax 3220 (br OH), 3030 (CH
aromatic), 2950 (CH aliphatic), 1620 (C=N), 1340 (C–N
agonists (Tully et al., 1991). Also, the pyrimidothiadiazi-
nes have been described as nucleoside analogues, (Haruo
et al., 1978) anti-inflammatory, hypotensive and diuretic
agents (Tatehiko et al., 1978a, b).
The biological activities of the previous compounds
persuaded us to study the antimicrobial and anti-inflam-
matory activities of selenolo[2,3-c]pyrazole and pyrim-
-
1 1
st), 720 (C–Cl) cm ; H NMR (d ppm, CDCl , 90 MHz):
3
2
.70 (3H, s, CH ), 7.20–7.60 (5H, m, ArH), 8.90 (1H, s,
3
0
0
ido[4 ,5 :4,5]selenolo[2,3-c]pyrazole derivatives.
1
3
CH=N), 10.80 (1H, s, OH); C NMR (d ppm, CDCl3,
00 MHz): 12.8 (CH pyrazole), 98.4 (C1), 118.3 (C6,
1
3
C10: Ph), 125.5 (C8 Ph), 126.8 (C7, C9: Ph), 136.1 (C11),
40.4 (C3), 143.5 (C2); Anal. Calcd. for C H ClN O
Materials and methods
Chemistry
1
1
1
10
3
(235.67): C, 56.06; H, 4.28; Cl; 15.04; N, 17.83. Found: C,
56.00; H, 4.35; Cl, 14.95; N, 18.00.
All melting points are uncorrected and measured on a
Fisher-John apparatus. IR spectra were recorded (KBr,
5-Chloro-3-methyl-1-phenyl-1H-pyrazole-4-carbonitrile
(3) A solution of oxime compound 2 (11 g, 0.047 mol) in
1
23

Med Chem Res
acetic anhydride (30 mL) was refluxed for 6 h. The solid
product which formed on pouring ice-water mixture was
recrystallized from ethanol as brownish-white crystals
139.6 (C3), 145.6 (C4), 145.8 (C6a), 169 (C14: CO); EIMS
?
m/z 320 [M ?1] (17), 319 [M] (100), 318 [M] -1 (9),
?
?
?
?
317 (M] -2 (26), 302 [M] -NH (27), 301 [M] -H O (16),
?
3
2
-
1
?
(
88.5 %, 9 g) yield; mp 118–120 °C. IR (KBr, cm ) mmax
274 [M] -HCONH (11); Anal. Calcd. for C H N OSe
2 13 12 4
3
030 (CH aromatic), 2950 (CH aliphatic), 2210 (C:N),
625 (C=N), 1340 (N–Ph st), 725 (C–Cl) cm ; H NMR
(319.23): C, 48.91; H, 3.79; N, 17.55. Found: C, 48.87; H,
3.70; N, 17.60.
-
1 1
1
(
d ppm, CDCl , 90 MHz): 2.50 (3H, s, CH ), 7.5 (5H, m,
3
3
1
3
4-(2-Chloroacetyl amino)-3-methyl-1-phenyl-1H-selenolo[2,3-
c]pyrazole-5-carboxamide (7) A solution of aminocarbox-
amide compound 6 (1 g, 0.003 mol) and chloroacetyl
chloride (0.40 mL, 3.5 mmol) in dioxane (30 mL) was
heated on water bath at 60–70 °C for 2 h. The solid product
which was obtained by cooling and pouring on diluted
sodium carbonate solution was recrystallized from ethanol
as white crystals in (81 %, 1.00 g) yield; mp: 250–252 °C;
ArH); C NMR (d ppm, CDCl , 100 MHz): 12.7 (CH
pyrazole), 95.2 (C1), 110.8 (C11), 118.3 (C6, C10 Ph),
3
3
1
1
6
3
25.4 (C8 Ph), 126.8 (C7, C9 Ph), 136.7 (C5), 141.2 (C3),
43.8 (C2); Anal. Calcd. for C H ClN (217.66): C,
8
1
1
3
0.70; H, 3.70; Cl, 16.29; N, 19.31. Found: C, 60.65; H,
.84; Cl, 16.35; N, 19.25.
2
-(4-Cyano-3-methyl-1-phenyl-1H-pyrazol-5-ylselanyl)ac-
-
1
etamide (5) This compound was prepared by adding (4 g,
.05 mol) of selenium metal to absolute ethanol (60 mL) in
IR (KBr, cm ) mmax 3380, 3300, 3180 (NH, NH ), 3050
2
0
(
CH aromatic), 2920, 2840 (CH aliphatic), 1680, 1630
1
a two-necked flask in an ice bath, and then, sodium boro-
hydride (3 g, 0.08 mol) was added in small portions till all
selenium metal dissolved. The reaction mixture was added
to (10.85 g, 0.05 mol) of chlorocyanopyrazole 3 with
stirring for 1 h., and then, the reaction mixture was refluxed
for 2 h followed by cooling at this stage, and the sodium
selenate salt 4 was formed. (4.70 g, 0.05 mol) of
chloroacetamide was added to the reaction mixture with
stirring and then was left overnight. The solid precipitate
which was formed by stirring and washing with water was
recrystallized from ethanol as white crystals (55 %, 11 g)
(2C=O amide), 1620 (C=N), 1345 (N–Ph st); H NMR (d
ppm, CDCl , 90 MHz): 2.50 (3H, s, CH ), 4.50 (2H, s,
3
3
CH ), 6.80 (s, 2H, NH ), 7.45–7.80 (5H, m, ArH), 10.40
2
2
1
3
(
(
(
(
1H, s, NH); CNMR (d ppm, CDCl , 100 MHz): 14.3
3
C7, CH pyrazole), 42.5 (C16, CH ), 98.1 (C3a), 117.8
3
2
C9, C13 Ph), 126.0 (C11 Ph), 127.2 (C10, C12 Ph), 130.4
C8), 136.1 (C5), 145.9 (C6a), 140.2 (C3), 164.7 (C15:
?
NHCO), 168.9 (C18: CONH ); EIMS m/z 395 [M] (100);
2
Anal. Calcd. for C H ClN O Se (395.71): C, 45.53; H,
1
5
13
4 2
3.31; Cl, 8.96; N, 14.16. Found: C, 45.60; H, 3.26; Cl, 9.10;
N, 14.25.
-
1
yield); mp 120–122 °C. IR (KBr, cm ) mmax 3450, 3350
NH ), 3030 (CH aromatic), 2920, 2850 (CH aliphatic),
0
0
5
-Chloromethyl-3-methyl-1-phenyl-pyrazolo[3 ,4 :5,4]se-
lenolo[2,3-e]pyrimidin-7(6H)-one (8): method A (3.20 g,
.01 mol) of chloroacetyl amino compound 7 in acetic
(
2
2
210 (CN), 1660 (CO amide), 1635 (N–H amide), 1620
1
0
(
C=N), 1335 (N–Ph st); H NMR (d ppm, CDCl3,
00 MHz): 2.50 (3H, s, CH ), 3.36 (2H, s, CH ), 7.14 (2H,
anhydride (20 mL) was refluxed for 2 h. The solid product
which formed on cooling was recrystallized from dioxane as
greenish white crystals in (78.5 %, 3 g) yield; mp:
4
3
2
1
3
s, NH disappeared by D O), 7.14–7.57 (5H, m, ArH);
2
C
2
NMR (d ppm, CDCl , 100 MHz): 13.8 (C6: CH pyrazole),
3
3
3
308–310 °C.
2.2 (C14: CH ), 100.5 (C4), 114.4 (C13: CN), 126.4 (C8,
2
C12 Ph), 129.5 (C10 Ph), 129.7 (C9, C11 Ph), 134.8 (C7),
39.1 (C3), 153.0 (C5), 169.8 (C15: CO). Anal. Calcd. for
C H N OSe (319.23): C, 48.90; H, 3.79; N, 17.55.
Method B A mixture of amino carboxamide compound 6
(3.32 g, 0.01 mol) and chloroacetyl chloride (3 mL,
1
1
3 12 4
0.038 mol) was heated on water bath at 60–70 °C for 3 h,
Found: C, 49.00; H, 3.65; N, 17.60.
-Amino-3-methyl-1-phenyl-1H-selenolo[2,3-c]pyrazole-5-
carboxamide (6) A mixture of selanyl compound 5
3.20 g, 0.01 mol) in sodium ethoxide was refluxed for
0 min. The solid product which formed on cooling was
recrystallized from dioxane as a white needle in (62.50 %,
then poured into cold water (100 mL), and neutralized with
sodium carbonate solution (10 %). The solid product
formed was recrystallized from dioxane. It was obtained as
a greenish white crystal in (3.0 g, 77 %) yield; mp:
4
(
-
1
1
308–310 °C; IR (KBr, cm ) mmax 3500–3400 (br NH),
3030 (CH aromatic), 2900, 2850 (CH aliphatic), 1645 (CO
-
1
1
amide), 1620 (C=N), 1340 (N–Ph st), 725 (C–Cl). H NMR
2
3
g) yield; mp 222–224 °C. IR (KBr, cm ) mmax 3380,
310, 3180 (2NH ), 3030 (CH aromatic), 1630 (CO
1
amide), 1590 (C=N), 1340 (N–Ph st); H NMR (d ppm,
DMSO-d , 400 MHz): 2.58 (3H, s, CH ), 6.82 (2H, s,
(d ppm, DMSO-d , 400 MHz): 2.60 (3H, s, CH ), 4.61
2
6
3
(2H, s, CH ), 7.35–7.70 (5H, m, ArH), 13.60 (1H, s, NH);
2
1
3
6
3
C NMR (d ppm, DMSO-d , 100 MHz), (100 MHz):
6
CONH ), 7.00 (2H, s, NH pyrazole), 7.22–7.65 (5H, m,
2
d = 13.2 (C9: CH pyrazole), 43.2 (C16: CH Cl), 94.0
2
3
2
1
3
ArH); CNMR (d ppm, DMSO-d , 100 MHz): 13.1 (C6:
(C3a), 118.1 (C11, C15 Ph), 122.0 (C13 Ph), 126.7 (C12,
C14 Ph), 126.8 (C10), 130.5 (C7a), 139.2 (C3), 146.4
(C8a), 150.6 (C5), 156.4 (C3b), 162.5 (C7: CO
6
CH pyrazole), 96.7 (C3a), 117.3 (C9, C13 Ph), 125.8
3
(
C11: Ph), 126.3 (C10, C12 Ph), 130.4 (C5), 136.6 (C8),
123

Med Chem Res
?
pyrimidine); EIMS m/z 377 [M] (100); Anal. Calcd. for
0
0
3-Methyl-1-phenyl-5-(p-tolylaminomethyl)-pyrazolo[3 ,4 :5,4]se-
lenolo[2,3-e]pyrimidin-7(6H)-one (9c): obtained by the reaction
with p-toluidine The solid product formed was recrystal-
lized from dioxane as white crystals in (80 %, 0.95 g)
C H ClN OSe (377.69): C, 47.70; H, 2.94; Cl, 9.39; N,
1
5
11
4
1
4.83. Found: C, 47.75; H, 3.00; Cl, 9.26; N, 15.00.
0
0
-1
3
-Methyl-1-phenyl-5-alkyl(arylamino)methyl-pyrazolo[3 ,4 :
yield; mp: 260–262 °C; IR (KBr, cm ) mmax 3400, 3350
(2NH st), 1665 (CO pyrimidine), 1620 (C=N), 1570 (N–H
5
,4]selenolo[2,3-e]pyrimidin-7(6H)-one (9a–f): general
procedure A mixture of chloromethyl compound 8 (1 g,
.65 mmol) and the corresponding amine (2.80 mmol) was
-
1
1
; H
amide deformation), 1345, 1290 (2N–Ph st) cm
2
NMR (d ppm, DMSO-d , 400 MHz): 2.13 (s, 3H, CH p-
6
3
refluxed in ethanol (20 mL) in the presence of few drops of
triethyl amine for 4 h. The solid product which separated
out during reflux was recrystallized from the proper
solvent.
tolyl), 2.61 (s, 3H, CH pyrazole), 4.30 (s, 2H, CH ), 5.90
3 2
(s, 1H, NH p-tolyl), 6.60–7.74 (m, 9H, ArH), 12.56 (s, 1H,
3
1
NH pyrimidine); C NMR (d ppm, DMSO-d , 100 MHz):
6
13.2 (C9: CH pyrazole), 20.5 (C24: CH p-tolyl), 46.6
3
3
(
C16: CH ), 97.9 (C3a), 113.4 (C19, C23: PhCH -p), 118.2
2 3
0
0
3
4
-Methyl-1-phenyl-5-phenylaminomethyl-pyrazolo[3 ,4 :5,
]selenolo[2,3-e]pyrimidin-7(6H)-one (9a): obtained by
(C21), 120.6 (C11, C15: Ph pyrazole), 125.8 (C13: Ph
pyrazole), 126.9 (C20, C22: PhCH -p), 129.8 (C12, C14:
3
the reaction with aniline The solid product formed was
Ph pyrazole), 130.6 (C10), 139.3 (C18: PhCH -p), 146.0
3
recrystallized from dioxane as white crystals in (0.90 g,
-
(C7a), 146.5 (C3), 147.1 (C8a), 151.0 (C5), 159.7 (C3b),
?
164.4 (C7: CO pyrimidine); EI-MS m/z 447 [M -1]
1
7
8 %) yield; mp: 288–289 °C; IR (KBr, cm ) mmax 3400,
3
350 (2NH st), 1650 (CO pyrimidine), 1620 (C=N), 1560
1
(53.6 %). Anal. Calcd. for C H N OSe (448.39): C,
22 19 5
(
N–H amide deformation), 1340, 1310 (2N–Ph st);
H
58.93; H, 4.27; N, 15.62. Found: C, 59.00; H, 4.35; N,
15.55.
NMR (d ppm, DMSO-d , 400 MHz): 2.60 (3H, s, CH ),
6
3
4
.34 (2H, s, CH ), 6.18 (1H, s, NHPh), 6.58–7.74 (10H, m,
2
0 0
-Methyl-1-phenyl-5-(p-anisylaminomethyl)pyrazolo[3 ,4 :5,4]se-
3
1
3
ArH), 12.61 (1H, s, NH pyrimidine); C NMR (d ppm,
DMSO-d , 100 MHz): 13.2 (C9: CH pyrazole), 43.9 (C16:
lenolo[2,3-e]pyrimidin-7(6H)-one (9d): obtained by the reaction
with p-anisidine The solid product formed was recrystal-
lized from dioxane as white crystals in (82 %, 1.01 g)
6
3
CH NH), 98.0 (C3a), 111.3 (C19, C23: NHPh), 116.8
2
(
C21: NHPh), 118.4 (C11, C15: Ph pyrazole), 126.1 (C13:
-
1
yield; mp: 238–240 °C; IR (KBr, cm ) mmax 3420 (br NH
st), 3350 (s NH st), 1660 (CO pyrimidine), 1625 (C=N),
Ph pyrazole), 127.2 (C20, C22), 127.8 (C12, C14 Ph
pyrazole), 130.5 (C10), 137.2 (C7a), 140.3 (C3), 140.6
1
565 (N–H amide deformation), 1340, 1300 (2N–Ph st); H
1
(
C18), 146.1 (C8a), 148.7 (C5), 153.2 (C3b), 162.0 (C7:
?
NMR (d ppm, CF CO D, 90 MHz): 3.30 (3H, s, CH
3
2
3
CO pyrimidine); EI-MS m/z 434 [M] (100 %). Anal.
Calcd. for C H N OSe (434.36): C, 58.07; H, 3.94; N,
pyrazole), 4.25 (3H, s, OCH ), 5.35 (2H, s, CH ),
3
2
13
C
2
1
17
5
7.35–7.90 (9H, m, p-sub anisyl, ArH pyrazole) ppm.
1
6.12. Found: C, 57.95; H, 4.00; N, 16.00.
NMR (d ppm, DMSO-d , 100 MHz): 13.3 (C9: CH
6
3
pyrazole), 24.6 (C25: CH p-anisyl), 44.9 (C16: CH ), 98.2
2
3
3
-Methyl-1-phenyl-5-(p-chlorophenylaminomethyl)-pyra-
(C3a), 114.5 (C19, C23: PhOCH -p), 119.3 (C21:
3
0
0
zolo[3 ,4 :5,4]selenolo[2,3-e] pyrimidin-7(6H)-one (9b):
obtained by the reaction with p-chloroaniline The solid
product formed was recrystallized from dioxane as white
crystals in (74 %, 0.92 g) yield; mp: 272–274 °C;IR (KBr,
cm ) mmax 3300, 3220 (2NH st), 1660 (CO pyrimidine),
1
PhOCH -p), 120.8 (C11, C15: Ph pyrazole), 126.1 (C13:
3
Ph pyrazole), 127.6 (C20, C22: PhOCH -p), 130.0(C12,
3
C14: Ph pyrazole), 130.7 (C10), 139.3 (C18: NHPhOCH3-
p), 146.8 (C7a), 146.8 (C3), 147.4 (C8a), 152.4 (C5), 158.1
-
1
(C3b), 164.9 (C7: CO pyrimidine); Anal. Calcd. for C22
625 (C=N), 1570 (N–H amide deformation), 1350, 1300
1
H N O Se (464.39): C, 56.90; H, 4.12; N, 15.08. Found:
19 5 2
(
2N–Ph st); H NMR (d ppm, DMSO-d , 90 MHz): 2.85
6
C, 56.84; H, 4.00; N, 14.95.
(
3H, s, CH pyrazole), 4.50 (2H, s, CH ), 6.95 (1H, s, NH
3
2
0
0
p-chlorophenyl), 7.30–7.85 (9H, m, ArH), 11.30 (1H, s,
3
3-Methyl-1-phenyl-(5-piperidin-1-ylmethyl)-pyrazolo[3 ,4 :5,4]se-
lenolo[2,3-e]pyrimidin-7(6H)-one (9e): obtained by the reaction
with piperidine The solid product formed was recrystal-
lized from ethanol as greenish white crystals in (81 %,
1
NH pyrimidine); C NMR (d ppm, DMSO-d , 100 MHz):
6
1
3.4 (C9: CH pyrazole), 48.0 (C16: CH NH), 98.2 (C3a),
3 2
1
12.8 (C19, C23: PhCl-p), 117.2 (C21), 118.4 (C11, C15:
-
1
Ph pyrazole), 126.2 (C13: Ph pyrazole), 127.3 (C20, C22:
0.86 g) yield; mp: 196–198 °C; IR (KBr, cm ) mmax 3480
(NH st), 3030 (CH aromatic), 2920, 2900, 2850 (CH ali-
phatic), 1650 (C=O pyrimidine), 1630 (C=N), 1560 (N–H
PhCl-p), 127.8 (C12, C14: Ph pyrazole), 130.4 (C10),
1
37.3 (C7a), 140.8 (C3), 142.6 (C18), 146.5 (C8a), 148.8
(
C5), 153.3 (C3b), 162.2 (C7: CO pyrimidine). EI-MS m/z
?
68 [M] (86 %). Anal. Calcd. for C H ClN OSe
amide deformation), 1345 (N–Ph), 1200 (C–N piperidine);
1
4
H NMR (d ppm, CDCl , 90 MHz): 1.35, 1.80 (6H, m,
2
1
16
5
3
(
468.81): C, 53.83; H, 3.44; Cl, 7.57; N, 14.94. Found: C,
4.00; H, 3.62;Cl, 7.62; N, 15.25.
(CH ) piperidine), 2.65 (3H, s, CH pyrazole), 2.80 (4H,
2
3
3
5
m, (CH ) N piperidine), 3.75 (2H, s, CH N), 7.45–7.90
2 2 2
1
23

Med Chem Res
1
3
-1
334–336 °C; IR (KBr, cm ) mmax 3050 (CH aromatic),
2920, 2850 (CH aliphatic), 1685 (CO pyrimidine), 1630
(
5H, m, ArH), 11.50 (1H, s, NH); C NMR (d ppm, CDCl3,
1
2
00 MHz) 12.8 (C9: CH pyrazole), 22.4 (C20: piperidine),
3
1
4.2 (C19, C21 piperidine), 40.8 (C18, C22 piperidine), 44.3
(C=N), 1345, 1310 (2N–Ph st); H NMR (d ppm, DMSO-
d , 400 MHz): 2.51 (3H, s, CH ), 5.54 (2H, s, CH N), 5.86
(
C16: CH N), 97.8 (C3a), 118.4 (C11, C15: Ph pyrazole),
2
6
3
2
1
3
1
1
12.6 (C13: Ph pyrazole), 126.5 (C12, C14:Ph pyrazole),
28.8 (C10: Ph pyrazole), 136.8 (C7a), 139.7 (C3), 144.9
(2H, s, NCH N imidazole), 6.98–7.72 (10H, m, ArH);
2
C
NMR (d ppm, DMSO-d , 100 MHz): 12.8 (C11: CH
6
3
(
C8a), 149.4 (C5), 152.3 (C3b), 163.4 (C7: CO pyrimidine).
pyrazole), 52.5, 58.8 (C5, C7: 2CH2 imidazole), 100.6
(C3a), 114.5 (C19, C23: Ph imidazole), 118.8 (C13, C17:
Ph pyrazole), 126.4 (C15: Ph pyrazole), 127.2 (C20, C22:
Ph imidazole), 127.9 (C21), 128.6 (C14, C16: Ph pyrazole),
Anal. Calcd. for C H N OSe (426.38): C, 56.34; H, 4.96;
20 21 5
N, 16.43. Found: C, 56.40; H, 5.00; N, 16.37.
0
0
3
-Methyl-1-phenyl-(5-morpholin-4-ylmethyl)-pyrazolo[3 ,4 :5,
]selenolo[2,3-e]pyrimidin-7(6H)-one (9f): obtained by the
1
1
31.0 (C12), 136.2 (C18: Ph imidazole), 138.5 (C9a),
4
42.0 (C3), 147.8 (C10a), 152.2 (C3b), 162.2 (C4a), 165.4
?
C9: CO pyrimidine). EI-MS m/z 446 [M] (55 %). Anal.
reaction with morpholine The solid product formed was
(
recrystallized from ethanol as green crystals in (80 %,
-
0
Calcd. for C H N OSe (446.37): C, 59.20; H, 3.84; N,
22 17 5
1
.85 g) yield; mp: 206–208 °C; IR (KBr, cm ) mmax 3450
NH st), 3020 (CH aromatic), 2950, 2900, 2850 (CH ali-
15.69. Found: C, 59.25; H, 3.78; N, 15.62.
(
phatic), 1645 (C=O pyrimidine), 1625 (C=N), 1560 (N–H
3-Methyl-1-phenyl-6-(p-chlorophenyl)-5,7-dihydroimidazo[3,4-
0
0
amide deformation), 1340 (N–Ph st), 1190 (C–N morpho-
1
b]pyrazolo[3 ,4 :5,4]selenolo [2,3-e]pyrimidin-9-one (10b)
The solid product formed by using method A was recrys-
tallized from chloroform as white crystals in (30 %,
line). H NMR (d ppm, CDCl , 90 MHz): 2.50 (3H, s,
3
CH ), 2.70–2.85 (4H, m, (CH ) N morpholine), 3.50–3.80
3 2 2
-
1
(
4H, m, (CH ) O morpholine), 3.90 (2H, s, CH N),
2
0.31 g) yield; mp: 328–330 °C; IR (KBr, cm ) mmax 3040
(CH aromatic), 2930, 2850 (CH aliphatic), 1680 (CO
2
2
1
3
7.20–7.80 (5H, m, ArH), 11.20 (s, 1H, NH); C NMR (d
ppm, CDCl , 100 MHz): 13.0 (C9: CH pyrazole), 51.3
1
pyrimidine), 1625 (C=N), 1350, 1315 (2N–Ph st); H NMR
3
3
(
C18, C22: (CH ) N morpholine), 55.8 (C16: CH ),
2
(d ppm, CF CO D, 90 MHz): 3.20 (3H, s, CH ), 5.20 (2H,
2
2
3
2
3
6
5.0(C19, C21: (CH ) O morpholine), 98.4 (C3a), 118.6
2
s, C–CH –N imidazole), 6.00 (2H, s, NCH N imidazole),
2 2
2
1
3
(
C11, C15: Ph), 122.4 (C13: Ph pyrazole), 126.8 (C12,
C14: Ph), 128.4 (C10), 136.6 (C7a), 139.7 (C3), 144.8
C8a), 149.0 (C5), 152.3 (C3b), 162.8 (C7: CO pyrim-
7.35–7.90 (9H, m, ArH); C NMR (d ppm, DMSO-d6,
100 MHz): 12.6 (C11, CH pyrazole), 52.6, 58.8 (C5, C7:
3
(
2CH imidazole), 101.3 (C3a), 113.4 (C19, C23: PhCl-p),
2
?
idine); EI-MS m/z 428[M] (100 %). Anal. Calcd. for
C H N O Se (428.36): C, 53.28; H, 4.47; N, 16.35.
118.6 (C13, C17: Ph pyrazole), 122.7 (C20, C22: PhCl-p),
126.5 (C15), 128.4 (C21: PhCl-p), 128.8 (C14, C16 Ph
pyrazole), 131.5 (C12), 138.3 (C18: PhCl-p), 139.0 (C9a),
1
9 19 5 2
Found: C, 53.35; H, 4.55; N, 16.46.
1
42.4 (C3), 146.9 (C10a), 153.0 (C3b), 162.8 (C4a), 166.4
0
3
-Methyl-1,6-diaryl-5,7-dihydroimidazo[3,4-b]pyrazolo[3 ,
(C9: CO). Anal. Calcd. for C H ClN OSe (480.82): C,
5
2
2
16
0
4
:5,4]selenolo[2,3-e]pyrimidin-9-one (10a–d)
54.96; H, 3.35; N, 14.57. Found: C, 55.10; H, 3.54; N,
14.65.
General procedure
3
-Methyl-1-phenyl-6-(p-tolyl)-5,7-dihydroimidazo[3,4-b]
0
0
Method A A mixture of N-aryl amino derivatives 9a–d
(
pyrazolo[3 ,4 :5,4]selenolo[2,3-e] pyrimidin-9-one (10c)
The solid product formed by using method B was recrys-
tallized from chloroform as white crystals in (58 %,
1 g, 2.2 mmol) was dissolved in warm ethanol (20 mL) and
then, formaldehyde (2 mL, 0.07 mol, 35 %) was added
dropwise with stirring for 15 min. The reaction mixture was
heated for 2 h at 50–60 °C. The solid product which sepa-
rated out during reflux was recrystallized from chloroform.
-
1
0.60 g) yield; mp: 308–310 °C; IR (KBr, cm ) mmax 3045
(CH aromatic), 2940, 2840 (CH aliphatic), 1675 (CO
1
pyrimidine), 1620 (C=N), 1340, 1310 (2N–Ph st); H-NMR
(
d ppm, DMSO-d , 400 MHz): 1.46 (3H, s, CH p-tolyl),
.25 (3H, s, CH pyrazole), 4.68 (2H, s, CH N imidazole),
3 2
6
3
Method B The N-aryl amino derivatives 9a–d (1 g,
2
2
.2 mmol) was dissolved in dioxane (20 mL), and then,
5
1
.47 (2H, s, NCH N imidazole), 6.55–7.66 (9H, m, ArH);
2
formaldehyde (2 mL, 0.07 mol, 35 %) was added dropwise
with stirring for 15 min. The reaction mixture was stirred
for additional 2 h at 50–60 °C. The solid product which
separated out on hot was recrystallized from chloroform.
3
C–NMR (d ppm, DMSO-d , 100 MHz): 12.7 (C11: CH
6
3
pyrazole), 22.5 (C24: CH p-tolyl), 52.4, 57.4 (C5, C7:
3
2
1
CH imidazole), 100.4 (C3a), 112.6 (C19, C23: PhMe-p),
2
19.6 (C13, C17 Ph pyrazole), 121.5 (C20, C22: PhMe-p),
0
3
-Methyl-1,6-diphenyl-5,7-dihydroimidazo[3,4-b]pyrazolo[3 ,
127.8 (C21: PhMe-p), 128.3 (C15), 129.8 (C14, C16 Ph
pyrazole), 130.0 (C12), 136.1 (C18 PhMe-p), 139.8 (C9a),
142.6 (C3), 145.3 (C10a), 152.6 (C3b), 162.3 (C4a), 165.9
0
4
:5,4]selenolo[2,3-e]pyrimidin-9-one (10a) The solid
product formed by using method B was recrystallized from
?
chloroform as white crystals in (34 %, 0.35 g) yield; mp:
(C9, CO pyrimidine); EI-MS m/z [M ? 1] 461 (100 %).
123

Med Chem Res
1
Anal. Calcd. for C H ClN OSe (460.40): C, 60.00; H,
2
pyrimidine), 1345, 1310 (2N–Ph); H NMR (d ppm,
DMSO-d , 90 MHz): 2.80 (3H, s, CH ), 5.50, 5.80 (4H, 2s,
3
19
5
4
.16; N, 15.21. Found: C, 60.10; H, 4.24; N, 15.32.
6
3
1
3
2
CH thiadiazine), 7.25–7.90 (10H, m, ArH); C–NMR (d
ppm, DMSO-d , 100 MHz): 14.1 (C12: CH pyrazole),
6
2
3
-Methyl-1-phenyl-6-(p-anisyl)-5,7-dihydroimidazo[3,4-b]
0
0
3
pyrazolo[3 ,4 :5,4]selenolo[2,3-e] pyrimidin-9-one (10d)
The solid product formed by using method B was recrys-
tallized from chloroform as white crystals (61 %, 0.60 g)
yield; mp: 312–314 °C; IR: 3030 (CH aromatic), 2950,
5
0.3, 64.4 (C6, C8: 2CH thiadiazine), 98.8 (C3a), 118.6
2
(
C22: Ph thiadiazine), 120.2 (C14, C18: Ph pyrazole),
1
21.6 (C20, C24: Ph thiadiazine), 123.4 (C21, C23: Ph
thiadiazine), 126.8 (C16: Ph pyrazole), 129.8 (C15, C17:
Ph pyrazole), 130.6 (C13: Ph pyrazole), 138.4 (C10a),
2
1
9
5
850 (CH aliphatic), 1670 (CO pyrimidine), 1625 (C=N),
1
340, 1310 (2N–Ph st). H NMR (d ppm, CF CO D,
3
2
1
1
40.2 (C19: Ph thiadiazine), 150.9 (C3b), 156.4 (C11a),
0 MHz): 3.00 (s, 3H, CH pyrazole), 3.90 (s, 3H, OCH ),
3
3
59.8 (C3), 162.3 (C4a), 165.6 (C10: CO pyrimidine); EI-
?
MS m/z: 478 [M] (100); Anal. Calcd. for: C H N OSSe
22 17 5
.20 (s, 2H, NCH imidazole), 6.00 (s, 2H, NCH N imi-
2
2
1
3
dazole), 7.25–7.90 (m, 9H, ArH). C–NMR (d ppm, DMSO-
d , 100 MHz): 13.2 (C11, CH pyrazole), 24.8 (C25: CH p-
(
478.44): C, 55.23; H, 3.58; N, 14.64; S, 6.70. Found: C,
6
3
3
55.30; H, 3.52; N, 14.71; S, 6.64.
anisyl), 55.6, 58.8 (C5, C7: 2CH imidazole), 102.3 (C3a),
2
0
0
1
1
14.8 (C19, C23: PhOMe-p), 120.4 (C13, C17 Ph pyrazole),
Ethyl (1-phenyl-3-methyl-7-oxopyrazolo[3 ,4 :5,4]selenolo
[2,3-e]pyrimidin-5-yl)acetate (13) A mixture of amino
carboxamide compound 6 (0.50 g, 0.0015 mol) and diethyl
malonate (1 mL, 6 mmol) in acetic acid (5 mL) was
refluxed for 1 h. The solid product which separated out on
hot was recrystallized from dioxane as brownish-white
needles in (67 %, 0.50 g) yield; mp 270–272 °C; IR (KBr,
22.2 (C20, C22: PhOMe-p), 127.8 (C21: PhOCH -p), 129.8
3
(
(
C15), 130.3 (C14, C16 Ph pyrazole), 131.5 (C12), 136.8
C18: PhOMe-p), 140.1 (C9a), 142.8 (C3), 146.2 (C10a),
153.6 (C3b), 162.8 (C4a), 166.2 (C9: CO pyrimidine); Anal.
Calcd. for C H N O Se (476.40): C, 57.99; H, 4.02; N,
2
3 19 5 2
1
4.70. Found: C, 58.00; H, 4.10; N, 14.75.
-
1
cm ) mmax 3150 (NH), 3030 (CH aromatic), 2920, 2850
CH aliphatic), 1740 (CO ester), 1650 (CO pyrimidine),
1
340 (N–Ph), 1150 (C-O st); H NMR (d ppm, DMSO-d ,
0
0
3
-Methyl-1-phenyl-7-oxopyrazolo[3 ,4 :5,4]selenolo[2,3-e]
(
pyrimidin-5-thione (11) A mixture of amino carboxamide
compound 6 (0.72 g, 0.002 mol) and carbon disulfide
1
4
2
6
00 MHz): 1.22–1.26 (3H, t, J = 12.58 Hz, CH ester),
3
(
2 mL, 0.026 mol) was refluxed on steam bath for 5 h. in
.51 (3H, s, CH pyrazole), 3.84 (2H, s, CH ), 4.16–4.20
3
2
the presence of anhydrous pyridine (2 mL). The solid
product which separated out on hot was recrystallized from
ethanol–dioxane mixture (3:1) as yellow crystals in
(
2H, q, J = 8.40 Hz, CH ester), 7.39–7.73 (5H, m, ArH),
2
1
3
1
2.81 (1H, s, NH); C NMR (d ppm, DMSO-d ,
00 MHz): 13.18 (C9: CH pyrazole), 14.51 (C19: CH
3
6
1
-
1
3
(
61.7 %, 0.5 g) yield; mp 340–342 °C; IR (KBr, cm
mmax 3300, 3100 (2NH), 3030 (CH aromatic), 2900, 2850
CH aliphatic), 1665 (CO pyrimidine), 1240 (C=S pyrim-
)
ester), 41.17 (C16: CH CO), 61.49 (C18: CH ester), 98.8
2
2
(
(
(
(
C3a), 118.18 (C11, C15: Ph), 120.94 (C13: Ph), 126.86
C12, C14: Ph), 130.54 (C10: Ph), 136.2 (C7a), 139.26
C3), 146.40 (C8a), 150.96 (C5), 155.09 (C3b), 159.57
C7: CO pyrimidine), 168.47 (CO ester); Anal. Calcd. for
(
1
idine), 1350 (N–Ph); H NMR (d ppm, DMSO-d6,
4
00 MHz): 2.50 (3H, s, CH ), 7.41–7.70 (5H, m, ArH),
3
1
3
12.76 (1H, s, NHCO), 13.00 (1H, s, NHCS); C NMR (d
ppm, DMSO-d , 100 MHz): 13.2 (C9: CH pyrazole), 66.8
C H N O Se (417.33): C, 51.81; H, 4.35; N, 13.43.
18 18 4 3
6
3
Found: C, 52.00; H, 4.22; N, 13.54.
Biological evaluation
(
C3a), 118.5 (C11, C15: Ph), 127.3 (C13: Ph), 130.6 (C12,
C14: Ph), 138.4 (C10: Ph), 139.8 (C7a), 142.1 (C3), 146.4
C8a), 152.9 (C3b), 192.9 (C7: CO pyrimidine), 198.4 (C5:
(
?
CS pyrimidine). EI-MS m/z 361 [M] (52.5). Anal. Calcd.
for C H N OSSe (361.29): C, 46.54; H, 2.79; N, 15.51;
Antimicrobial activity
1
4 10 4
S, 8.87. Found: C, 46.44; H, 2.85; N, 15.40; S, 8.90.
The fungal and bacterial strains are obtained from the
culture collection of Assiut University Mycological Center
(AUMC). The fungal strains were grown in sterilized 9-cm
Petri dishes containing Sabouraud’s dextrose agar (SDA)
supplemented with 0.05 % chloramphenicol to suppress
bacterial contamination (Al-Doory 1980). From these cul-
tures, agar disks (10 mm diameter) containing spores and
hyphae were transferred aseptically to screw-topped vials
containing 20 mL sterile distilled water. After thorough
shaking, 1-mL samples of the spore suspension were
pipetted into sterile Petri dishes, followed by the addition
00
00
0
0
0
0
1
,7-Diphenyl-3-methylpyrazolo[3 ,4 :5 ,4 ]selenolo[2 ,3 :5,
]pyrimido[2,3-b][1,3,5]thiadiazin-10(9H)-one (12)
mixture of the thione compound 11 (0.36 g, 0.001 mol),
6
A
formaldehyde (2 mL, 0.07 mol, 35 %) and aniline
(
0.2 mL, 0.002 mol) was refluxed in ethanol (20 mL) in
the presence of few drops of acetic acid (0.2 mL) for
0 min. The solid precipitate which separated out on hot
was recrystallized from dioxane as yellow crystals in
3
-
1
(
55 %, 0.30 g) yield; mp: [360 °C. IR (KBr, cm ) mmax
030 (CH aromatic), 2920, 2820 (CH aliphatic), 1660 (CO
3
1
23

Med Chem Res
123

Med Chem Res
1
23

Med Chem Res
Scheme 1 Synthesis of
aminoselenolopyrazole
carboxamide 6. Reagents and
1
3
OH
6
2
13
6
CHO
Cl
3
4
N
N
4
3
conditions: a H
EtOH; b Ac O, reflux; c Se,
NaBH , EtOH;
2
NOH, AcONa,
1
N
2
1
2
N
N
b
N
N7
5
Cl
N7 5 Cl
4
a
e
2 2
d ClCH CONH ; e EtONa,
EtOH
8
1
2
12
1
8
1
1
9
1
9
1
0
1
0
1
2
3
7
NH2
6
13
4
c
3
3a
NH2
4
N
3
4
d
1
N
2
1
4
1
5
N
15
Se
2
N
1
NH2
O
N8 6a
Se
6
N7 5
N
SeNa
O
N
13
9
12
8
9
1
0
11
12
11
10
6
5
4
of 15 mL liquefied SDA medium which was then left to
solidify. The bacterial strains were grown on nutrient agar
caliper (SMIEC, China). The tested compounds 9a–e at
doses 10 mg/kg of body weight were injected to five
groups of rats, while the last group was treated with
indomethacin drug as a positive control. Paw thickness was
measured just before and after the carrageenan injection
(negative control). The difference between the thicknesses
of the two paws was taken as an indication of edema and
measured at 0.5, 1, 2, 3, 4 and 5 h after injection of the
tested compounds.
(
NA). The tested compounds (5, 6, 7, 8 and 9a–f) and
reference compound (levofloxacin) were dissolved in
DMSO to give 2.0 % concentration. The specific names
and numbers are indicated in Tables 1 and 2. Only 3 bac-
terial strains were kindly provided by the Microbiology
Department, Faculty of Medicine, Assiut University.
Antifungal and antibacterial activities were determined
according to the method reported by Kwon-Chung and
Bennett (Kwon-Chung and Bennett 1992) using 5-mm-di-
ameter filter wells loaded with 50 lL of the solution under
investigation (2.0 %), and the inoculated plates were
incubated at 30 °C.
Results and discussion
Chemistry
Anti-inflammatory activity
The synthetic methods for target compound 6 are illus-
trated in Scheme 1. Reaction of 5-chloro-3-methyl-1-phe-
nyl-1H-pyrazole-4-carbaldehyde (1) with hydroxyl amine
hydrochloride in ethanol in the presence of sodium acetate
afforded the corresponding oxime 2. The latter was dehy-
drated upon heating with acetic anhydride into the corre-
sponding pyrazole carbonitrile derivative 3. The latter
compound was elucidated on the basis of TLC, m.p. and IR
spectrum which revealed the disappearance of absorption
Anti-inflammatory activity screening for the synthesized
compounds 9a–e was measured in vivo using carrageenan-
induced rat paw edema assay in comparison with indo-
methacin as reference drug (Winter et al., 1962; Adeyemi
et al., 2002). The test is based on the pedal inflammation in
rat paws induced by sub-plantar injection of 100 lL of 1 %
freshly prepared solution of carrageenan in distilled water
into the right hind paws of each rat of all the groups; the
tested compounds were dissolved in distilled water with
sonication. Male adult albino rats (120–150 g) were divi-
ded into six groups; each group contains three animals. The
thickness of the rat paw edema was measured by a Vernier
-
1
band at 3320 cm characteristic for OH of oxime and
-
1
appearance of band at 2210 cm for CN group. Incorpo-
rating of selenium metal fused to pyrazole ring was
achieved through reduction of selenium by sodium boro-
hydride in ethanol to afford the intermediate sodium salt 4
123

Med Chem Res
1
3
which formed in situ and not isolated. The selanyl acet-
amide 5 was formed by adding chloroacetamide to the
sodium selenate salt 4 in situ. The selanyl acetamido
compound underwent Thorpe–Ziegler cyclization upon
heating with ethanolic sodium ethoxide to give 4-amino
selenolo[2,3-c]pyrazole carboxamide (6). The latter com-
for (NH) group. C NMR in CDCl showed signals at d 22.4,
3
24.2 and 40.8 characteristic for aliphatic carbons of piperidine
ringin additionto the signal at d 163.4 characteristic for CO of
pyrimidine.
Treatment of compounds 9a–d with formaldehyde in
refluxed ethanol or by stirring in dioxane under Mannich
conditions afforded imidazopyrimidoselenolopyrazole
derivatives 10a–d. Compounds 10a–d were confirmed on
the basis of analytical and spectral data. IR spectrum of
compound 10a revealed the disappearance of absorption
1
pound was established by IR, H NMR and mass spectra. IR
-1
revealed disappearance of absorption band at 2210 cm
for CN group in compound 6 and appearance of absorption
-
1
bands at 3380, 3310 and 3180 cm characteristic for 2NH
2
-1
1
for CO amide. H NMR in
-1
groups and at 1630 cm
bands at 3400, 3350 cm characteristic for 2NH groups
-
and appearance of absorption bands at 1685 cm for C=O.
1
DMSO-d showed two signals at d 6.80 and 7.00 ppm for
6
1
3
1
2
NH groups. C NMR showed signal at d 169 charac-
H NMR spectrum of compounds 10a in DMSO-d showed
6
2
teristic for CO amidic group. Mass spectrum of compound
showed a peak at 319.77 (100 %) as a molecular ion peak
two singlet signals at d 5.54 and 5.86 ppm characteristic
for 2 CH imidazole groups. C NMR of compound 10a in
1
3
6
and base peak (Scheme 1).
2
DMSO-d showed two signals at d 52.5 and 58.8 charac-
6
Reaction of 6 with chloroacetyl chloride in dioxane fol-
lowed by neutralization with sodium carbonate solution
afforded the chloroacetyl amino 7, which underwent ring
closure using acetic anhydride to afford the chloromethyl
pyrimido compound 8. The latter compound can be obtained
by heating the amino carboxamide 6 with excess chloroacetyl
chloride upon heating with steam bath followed by neutral-
izing the reaction mixture using diluted sodium carbonate
solution. The structure of the produced compound 8 was
teristic for 2 CH groups of imidazole and signal at d 165.4
for CO pyrimidine (Scheme 2).
2
Heating of compound 6 with carbon disulfide in pyridine
on steam bath for 10 h afforded the pyrimidine thione 11
which underwent cyclization under double Mannich
conditions using formaldehyde and aniline in refluxed
ethanol in the presence of few drops of acetic acid to
give a new heterocyclic system namely 1,7-diphenyl-3-
0
0
00
0
0
0
0
methyl[1,3,5]thiadiazino[2 ,3 :2 ,1 ]pyrimido[4 ,5 :4,5]se-
lenolo[2,3-c]pyrazole-10(9H)-one 12. The structure of
compound 12 was established by TLC, mp, elemental and
spectral data. IR spectrum showed absorption bands at
1
elucidated on the basis of IR, H NMR and mass spectra. IR
spectrum revealed the disappearance of absorption bands at
-
1
3
380, 3300, 3180 cm for (NH, NH ) in compound 7 and
2
-1
-1
1
appearance of broad absorption band at 3500–3400 cm for
1
NH) and absorption band at 1645 cm for (CO). H NMR
1660 cm for (CO pyrimidine). H NMR spectrum in
-1
(
(DMSO-d ) showed two singlet signals at d 5.50 and
6
spectrum of compound 8 in (DMSO-d ) revealed appearance
6
5.80 ppm characteristic for 2CH groups. Mass spectrum of
2
of signals at d 4.61 ppm characteristic for CH and at d
compound 12 showed a peak at 478.00 as a molecular ion
peak and a base peak. The reaction of compound 6 with
diethyl malonate in acetic acid afforded Schiff’s base fol-
lowed by elimination of ethanol to give pyrimidoselenolo-
pyrazole acetic acid ethyl ester compound 13. Formation of
compound 13 was established on the basis of spectral
analysis. IR spectrum showed absorption bands at
2
1
3
1
3.60 ppm for NH. C NMR showed signal at d 162.5 ppm
characteristic for CO group of pyrimidine. Also compound 8
was confirmed by mass spectrum which showed a molecular
ion peak at (m/z) 377.40 (100 %) which was in agreement
with the postulated structure.
The chloromethyl derivative 8 underwent nucleophilic
substitution reactions with various primary and secondary
amines in refluxing ethanol to afford the alkyl(aryl)
aminomethylpyrimido derivatives 9a–f. Formation of com-
pounds 9a–f was established by elemental and spectral anal-
ysis. IR spectrum of compound 9c showed two absorption
-1
-1
3150 cm for (NH) and at 1740, 1650 cm for CO ester
1
and CO amide, respectively. HNMR in (DMSO-d6)
showed triplet and quartet signals at d 1.22–1.26 and
1
3
C
4.16–4.20 ppm characteristic for CH CH ester group.
2
3
NMR showed signals at 159.57 and 168.47 for C=O amide
and C=O ester, respectively (Scheme 3).
-1
bands at 3400, 3350 cm according to 2NH groups, while
-1
compound 9e showed absorption band at 3480 cm for one
1
NH group. H NMR spectrum of compound 9c showed two
The suggested mechanism for proving structure of
compound 12 contains the following steps. The first step
includes condensation between two molecules of aniline
and formaldehyde to afford the corresponding Schiff’s base
a. Next, a nucleophilic attack of the thiole group of the
tautomerized form of compound 11 to the imino group of
the Schiff’s base a was carried out to afford intermediate
c which underwent second nucleophilic addition of NH to a
second molecule of formaldehyde to give intermediate
singlet signals at d 2.13 and 2.61 ppm for 2 CH groups of p-
3
tolyl and pyrazole rings, respectively, multiplet signals at d
6
.60–7.74 ppm characteristic for aromatic protons and singlet
1
signal at d 12.56 ppm for NH group, while H NMR of
compounds 9e in (CDCl ) showed multiplet signals at d 1.35,
3
1
.80 and 2.80 ppm characteristic for aliphatic protons of
piperidinyl group and showed singlet signal at d 11.50 ppm
1
23

Med Chem Res
Scheme 2 Chloroacetylation of
amino carboxamide followed by
nucleophilic substitution to
amines. Reagents and
17
1
6
Cl
1
4 15
7
4HN
3
3
a
O
18
conditions:
19
NH2
2
a ClCH
reflux; c ClCH
d R NH/EtOH;
e HCHO/dioxane, stirring
2
COCl/dioxane; b Ac
2
O,
N
1
5
Se
6
2
COCl, fusion;
N
6
a
a
O
R
1 2
8
6
13
9
12
1
0
11
c
b
7
9
16
4
5
R1
9
1
6
3
4
N
N
N
5
2
3a 3b
3
Cl
R2
N1
N
3
a 3b
d
NH
7
6
2
N 1
N
NH 6
7
7a
8
9 a-f
1
5
Se
a
8
10
7a
O
8
a
1
5
10
Se
8
8
O
14
11
1
1
14
12
1
3
9 a-d
e
1
2
1
3
1
1
9
9
9
9
9
9
a: R = Ph, R = H
1 2
5
4
N
3
4a
6
b: R = PhCl-p, R = H
1
2
2
3a 3b
c: R = PhMe-p, R = H
2
N 1
N
N R
1
2
d: R = PhOMe-p, R = H
1
9
N
7
1
2
1
7
e: R ,R = -(CH )
2 5
f: R , R = -(CH ) O
2 4
8
1
2
10a Se
9a
10
16
13
14
1
2
O
1
0 a-d
1
5
1
0a: R= Ph
0b: R= PhCl-p
0c: R= PhMe-p
0d: R= PhOMe-p
1
1
1
Scheme 3 Reaction of amino
carboxamide 6 with carbon
disulfide followed by double
Mannich reaction. Reagents and
9
1
6
18
9
4
4
N
17
3
19
5
3
H
5
S
3a
3
b
N
2
2
3
a
3b
N 1
N
c
a
O
N 1
NH6
7
6
conditions: a CS
b HCHO, PhNH
c CH (CO Et) /AcOH
2
/pyridine;
NH 6
7
N
/EtOH, AcOH;
15
8a
2
Se 7a
15
8a
7
a
10
10
Se
8
8
O
2
2
2
14
O
1
1
11
14
11
13
12
13
13
12
b
1
2
5
S
4
N
4
a
6
3
20
7
21
22
3
a 3b
19
24
2
N
N
1
N
0
1
N
9
8
11a
10a
18
1
3
Se
23
11
O
17
14
1
2
15
16
alcohol d. The latter intermediate d losses water molecule
in the presence of acidic medium to form the corresponding
carbocation e. Finally, the lone pair of unshared electrons
of NH group of pyrimidine attacks the carbocation with
loss of a proton to afford the thiadiazino compound 12. The
suggested mechanism was described (Scheme 4).
123

Med Chem Res
H
H
activity. The p-chlorophenyl 9c and p-anisyl 9d derivatives
showed the highest antibacterial activity. Compound 9c
showed high antibacterial activity against all tested gram
H
H
C O + PhNH
2
C N Ph
a
H
N
S
SH
+
?
ve strains (18–24 mm), while compound 9d showed high
N
H
H
N
N
antibacterial activity against most tested gram –ve bacteria,
especially Escherichia coli (20 mm) and Klebsiella
(18 mm) and higher than the reference drug (17 and
C N Ph
NH
NH
N
N
Se
Se
b
Ph
Ph
O
O
1
1
14 mm), respectively. On the other hand, the phenyl and p-
S
S
chlorophenyl derivatives 9a, 9b showed similar interme-
diate antibacterial activities against gram ?ve bacteria
N
N
N
N
N
H N. .
NH
Ph
NH
Ph
N
N
(10–14 mm), while the piperidinyl and morpholinyl
Ph
Se
Ph
Se
c
OH
derivatives 9e, 9f are resistant to all strains of bacteria
(\10 mm).
O
N
-H
2
O
O
H
H
d
C O
H+
In a similar manner, the pyrazolylselanyl acetamide 5
showed almost high antifungal activity, especially against
Candida albicans (16 mm). On the other hand, the activity
decreased in amino carboxamide compound 6 was found to
be intermediate against Candida albicans (12 mm) and
resistant to the rest of antifungal strains (\10 mm). In
contrast to antibacterial activity, the piperidinyl and mor-
pholinyl derivatives 9e, 9f showed higher antifungal activ-
ities than the other aryl amino derivatives 9a–d. Compound
S
S
N
+
N
N
N
.
.
N
N
NH
Ph
N
NH
Ph
+
CH2
H C
2
Ph
Se
Ph
Se
O
O
f +
-H
e
S
N
N
N
N
Ph
N
Ph
Se
9e showed high antifungal activity against Candida albicans
O
12
(18 mm), Fusarium oxysporum (20 mm), Scopulariopsis
brevicaulis (20 mm) and Geotrichum candidum (18 mm),
while compound 9f showed high antifungal activity against
Candida albicans (17 mm), Trichophyton rubrum (28 mm)
and Geotrichum candidum (22 mm).
Scheme 4 The suggested reaction mechanism for formation of
thiadiazinopyrimidoselenolopyrazole 12
Pharmacology
Anti-inflammatory
Antimicrobial activity
Statistical analysis
Structure–Action Relationship (SAR) of the tested com-
pounds Some tested compounds showed high antibacterial
and antifungal activities. In case of antibacterial activity,
we found that pyrazolyl selanyl acetamide 5 showed high
antibacterial activity against some strains of gram (-ve)
bacteria such as Esherichia coli and Klebsiella, and its
values (24 and 22 mm) are higher than levofloxacin (17
and 14 mm) and moderate activity against Pseudomonas
aeruginosa (14). Ring closure of selanylacetamide 5 to
afford amino carboxamide 6 decreases the antibacterial
activity against gram (-ve) strains in spite of increasing
activity against gram (?ve) bacteria such as Clostridium
The results were analyzed by one-way analysis of variance
(ANOVA) followed by Newman–Keuls multiple compar-
ison test as a post-test. These analyses were carried out
using computer Prism program for Windows version 3.0
(GraphPad software, Inc, San Diago CA, USA). The sig-
nificant difference between groups was accepted at
P \ 0.05 *, 0.01** or 0.001 ***
The following Table 3 represented decrease in the
thickness of rat paw edema (mm) by the effect of the tested
compounds with time compared to the reference drug
(indomethacin).
(
16 mm). The chloroacetyl amino selenolopyrazole 7
A plot represented the decrease in thickness of paw
edema inhibition in rats by the effect of indomethacin and
the tested compound with time is described in Fig. 1.
The inflammatory response is represented as the time
course of the percentage of the increase in paw swelling as
the area under the curve response and as the percentage of
paw edema inhibition. Calculation for each compound at
each time point was done using the following ratio:
showed moderate antibacterial activity against most strains
of bacteria (10–12 mm), while the antibacterial activity of
chloromethyl pyrimido compound 8 increased especially
against Clostridium in which its value (20 mm) is higher
than the reference drug (15 mm). Substitution of chlorine
atom in chloromethyl compound 8 with nucleophilic pri-
mary and secondary amines strongly affected antibacterial
1
23

Med Chem Res
Table 3 Effect of pyrazoloselenolopyrimidine derivatives (9a–e) on carrageenan-induced paw edema in rats (zone of inhibition in mm)
Compound The thickness of rat paw edema (mm)
Time (h) Negative
control
Indomethacin
(0.028 mmol)
9a
(0.023 mmol)
9b
(0.021 mmol)
9c 9d
(0.022 mmol) (0.022 mmol)
9e
(0.023 mmol)
0
1
2
3
4
5
.5
0.717 ± 0.016
0.683 ± 0.016
0.683 ± 0.016
0.683 ± 0.016 0.700 ± 0.0
0.700 ± 0.0
0.700 ± 0.0
0.717 ± 0.016 0.5500 ± 0.028
0.683 ± 0.016 0.6333 ± 0.044 0.650 ± 0.028 0.700 ± 0.0
0.650 ± 0.028
0.717 ± 0.016
0.717 ± 0.016
0.717 ± 0.016
0.717 ± 0.016
0.483 ± 0.016
0.450 ± 0.028
0.417 ± 0.016
0.383 ± 0.016
0.683 ± 0.016
0.617 ± 0.016
0.483 ± 0.016
0.483 ± 0.016
0.550 ± 0.028 0.633 ± 0.016 0.683 ± 0.016 0.617 ± 0.016
0.483 ± 0.016 0.583 ± 0.016 0.683 ± 0.016 0.567 ± 0.044
0.467 ± 0.016 0.550 ± 0.028 0.667 ± 0.016 0.550 ± 0.028
0.400 ± 0.0
0.483 ± 0.016 0.633 ± 0.016 0.500 ± 0.0
0
0
0
0
.75
*
*
** ***
**
***
*
**
*
***
Negative control
Indomethacin
9a
***
**
*
* *
*
*
*
*
*
*
***
*
***
.50
.25
.00
*
9
9
9
9
b
c
d
e
0
.5
2
3
4
5
1
Ti me (h)
Fig. 1 A diagram represents decrease in thickness of rat paw edema of compounds 9a–e compared with indomethacin for 5 h
edema is the same at the control time and, after 30 min, has
Percentage of edema inhibition ð%Þ
no significant difference in the size of edema between
¼
where V is the volume of paw edema in negative control
ðV À V = V Þ Â 100
ð1Þ
c
t
c
indomethacin and the tested compounds. As shown in
Tables 4, 5 and Fig. 2, the anti-inflammatory activity of
p-chlorophenyl compound 9b is higher by 1.43 times than
indomethacin after 30 min, while a significant difference
exists between indomethacin and p-anisyl 9d which has
P value less than 0.05 that means compound 9d does not
affect inflammation of rats after 1 h.
c
just after carrageenan injection and V . is the volume of
t
paw edema in treated group. The following Table 4 and
Fig. 2 represented increase in the percentage of paw edema
inhibition for the tested compounds compared to the
indomethacin.
Potency of the tested compounds was calculated relative
to indomethacin ‘‘reference standard’’ treated group
according to the following equation:
After 2 h, all compounds showed significant differences
from indomethacin as p-chlorophenyl amino compound 9b
Percentage edema inhibition of tested compound treated group
Percentage edema inhibition of indomethacin treated group
Potency ¼
ð2Þ
From the previous results, we found that some of the
tested compounds showed high anti-inflammatory activity
compared with indomethacin. For example, the size of
showed the lowest significance from the other tested
compound, which means compound 9b is similar in its
effect to indomethacin. After 3 h., the p-chlorophenyl
123

Med Chem Res
Table 4 The increase in percentage of edema inhibition (%) of compounds 9a–e with time (h)
Time (h)
Paw edema inhibition (%)
Indomethacin
9a
4.74
9b
6.82
9c
2.37
9d
2.37
9e
0
1
2
3
4
5
.5
4.74
23.29
32.64
37.23
41.84
46.58
2.37
9.34
4.74
4.74
13.6
9.34
11.72
18.69
23.29
32.64
2.37
4.74
24.97
34.1
13.95
20.92
23.29
30.26
13.94
32.77
32.77
4.74
36.29
45.43
6.97
11.72
Fig. 2 A diagram represents
the increase in percentage of
edema inhibition (%)
50
4
3
2
1
0
0
0
0
0
Indomethacin
compounds 9a–e with time (h)
9
a
9
b
9
9
c
d
9
e
0.5
1
2
3
4
5
Time (h)
Table 5 The potency of compounds 9a–e relative to indomethacin
Time (h) Potency relative to indomethacin
9
a
9b
9c
9d
9e
0
1
2
3
4
5
.5
1
1.43
0.58
0.76
0.91
0.86
0.97
0.50
0.40
0.36
0.50
0.56
0.70
0.50
0.10
0.14
0.12
0.17
0.25
0.50
0.40
0.42
0.56
0.56
0.65
0.20
0.14
0.37
0.78
0.70
amino compound 9b showed no significance from indo-
methacin, while the p-tolyl amino derivative 9c and the
piperidinyl compound 9e showed low significance
imidazopyrimidine and thiadiazinopyrimidine. Some of
the synthesized compounds were screened against some
strains of bacteria and fungi. The antimicrobial activities
results demonstrated that the most of the tested com-
pounds showed antibacterial and antifungal activities. On
the other hand, the aryl aminomethyl pyrimidoselenolo
pyrazolone derivatives especially compounds 9b then 9a
showed high anti-inflammatory activity compared with
indomethacin. Consequently, this degree of biological
activity displayed by the novel synthesized compounds
encourages us to achieve more active derivatives in the
ongoing studies.
(
P \ 0.05). After 4 h., the phenyl amino 9a and p-chlor-
ophenyl amino 9b showed low significant difference from
the reference compound (P \ 0.05). After 5 h., the p-
chlorophenyl amino 9b showed no significance which
means that compound 9b has the highest anti-inflammatory
effect of the tested compounds.
In summary, the present work focused on the syn-
thesis of new selenolo[2,3-c]pyrazole compounds fused
to other heterocyclic rings namely pyrimidine,
1
23

Med Chem Res
Conclusion: the antibacterial, antifungal and anti-
inflammatory activity of novel selenolo[2,3-
c]pyrazoles
derivatives and a substituted azabicyclo[3.1.0]hexane-2,4-dione.
Anticancer Res 21:2313–2321
Brown ML, Cheung M, Dickerson SH, Drewry DH, Lackey KE, Peat
AJ, Thomson SA, Veal JM, Wilson JLR (2004) PCT Int. Appl.
WO 2004, 9596. Chem Abstr 140:128436y
In this work, we synthesized the amino selenolo[2,3-
c]pyrazole carboxamide compound 6 by an innovative
method and used it as a versatile precursor for synthesis of
other new heterocyclic rings fused to selenolopyrazole
moiety. Some of the newly synthesized compounds were
tested against some strains of bacteria and fungi in addition
to examining the anti-inflammatory activity for these
compounds in which some of them showed promising
results as antibacterial, antifungal and anti-inflammatory
agents.
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Acknowledgments The authors are very grateful to Prof Dr./Kamal
I. Ali, the chairman of Chemistry Department, for all facilities pro-
vided to us and very grateful to Prof. Dr/Alaa Arafat, Professor of
Pharmaceutical Organic Chemistry, Faculty of Pharmacy and all the
staff members of Pharmacology Department and Bacteriology
Departments, Faculty of Medicine, Assiut University for performing
the anti-inflammatory and some of the antimicrobial evaluation. The
authors are very grateful to staff members at Assiut University
Mycological Center for performing the antifungal activity and some
of the antibacterial activity.
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