Synthesis and biological evaluations of new nitric oxide-anti-inflammatory drug hybrids

Article abstract of DOI:10.1016/j.bmcl.2017.08.023

Three novel series of nitroso derivatives (11–15), isoxazolopyrazoles (17a–c) and isoxazolo[3,4-d]pyridazines (18a–c) were prepared from the hydroxyimoyl chloride 10. In vitro COX1?2 inhibition activities were evaluated, both of 17b and 18a proved a promi

Full text of DOI:10.1016/j.bmcl.2017.08.023

Accepted Manuscript
Synthesis and biological evaluations of new nitric oxide -anti-inflammatory
drug hybrids.
Eman K.A. Abdelall, Abdou O. Abdelhamid
PII:
S0960-894X(17)30824-7
DOI:
http://dx.doi.org/10.1016/j.bmcl.2017.08.023
BMCL 25217
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
8 July 2017
8 August 2017
11 August 2017
Please cite this article as: Abdelall, E.K.A., Abdelhamid, A.O., Synthesis and biological evaluations of new nitric
oxide -anti-inflammatory drug hybrids., Bioorganic & Medicinal Chemistry Letters (2017), doi: http://dx.doi.org/
10.1016/j.bmcl.2017.08.023
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Synthesis and biological evaluations of new nitric oxide -anti-inflammatory drug hybrids.
Eman K. A. Abdelall *a, Abdou O. Abdelhamidb
*aDepartment of Pharmaceutical Organic Chemistry, Faculty of Pharmacy, Beni-Suef
University, Beni-Suef, 62514, Egypt.
bDepartment of Chemistry, Faculty of Science, Cairo University, Giza 12613, Egypt.
___________________________________________________________________________
Abstract ― Three novel series of nitroso derivatives (11-15), isoxazolopyrazoles (17a-c) and
isoxazolo[3,4-d]pyridazines (18a-c) were prepared from the hydroxyimoyl chloride 10. In vitro
COX1\2 inhibition activities were evaluated, both of 17b and 18a proved a promising inhibitory
activity with IC50 = 1.12, 0.78 µM in sequent. Carragenan induced Paw edema, ulcer liability,
nitric oxide (NO.) release and histopathological study were determined. Most of the prepared
compounds showed excellent activities. Reactions of 2-aminopyridine and enaminone with
hydroxyimoyl chloride 10 were investigated and proved by 2D NMR. Molecular docking for
most active compounds was operated giving a hint for compound –receptor interactions.
_____________________________________________________________________________
(NSAIDS) Nonsteroidal anti-inflammatory drugs are usually recommended during the
medical treatments of various diseases. The use of classical non-steroidal anti-inflammatory
1
drugs is commonly with the incidence of gastric ulcer and bleeding . This classical type as
aspirin or indomethacin is famously known to exert the anti-inflammatory effects through
inhibition of both constitutive COX-1 and induced COX-2 but in a non selective manner.
2
Recently, the discovery of celecoxib (celebrexTM) (1) , valdecoxib (bextraTM)
(2) and
rofecoxib (vioxxTM) (3), as Coxibs candidates (Fig.1), confirms the concept of preparation of
more selective COX-2 inhibitors with a minimal gastric effect. Unfortunately, valdecoxib (2)

2
3
and rofecoxib (3) are withdrawn from the market due to cardiovascular complications. Nitric
oxide (NO.) is a biologically active molecule, known to play a major role in some pathological
conditions as vasodilatation of blood vessels, inhibition of platelet aggregation, and gastric
mucosal healing 4,5. So, NSAIDs with a (NO.) donor group have been developed as a new trend.
Such trend maintains the effective treatment of inflammation with minimum gastric and
cardiovascular side effects 6,7. Several NSAIDs-NO drug hybrids 4 8, 5 9 or NMI-1093 (6) 10 in
literature achieved such target for the appearance of both cardio and gastric safe anti-
inflammatory drugs. These NO-NSAIDs hybrids belonged to major four groups: 1- C-NO -
NSAID to which our research is directed, 2- O-NO-NSAIDS as nitro-esters, 3- N-NO-releasing
for which diazoniumdioleate is related and 4- S-NO- NSAIDS known as a sulfohydroxamic acid
9
ester group . Moreover, hydroximoyl chloride reagents as nitrile oxide precursors which
formed in-situ via removal of 1mole HCl. Such nitrile oxides being a versatile intermediate to
get in many 1,3-dipolar regio- specific or face-selective cycloadditions reactions. These nitrile
oxides could react with a diverse of reagents which belonged to either ח -deficient or ח -
excessive types as aminopyridines, o-phenylenes, aminophenols, and enaminones resulting in a
variety of heterocycles with a proper nitric oxide donor moiety (N-O) 11,12. A close sight to
celecoxib as a selective COX-2 and the NSAIDs-NO. hybrid drugs 4 or 5, all are diaryls with a
pyrazole central ring. As a continuation of previous work 13-18, the aim research is directed
toward the preparation of new diaryls with a central pyrazole ring and one of these aryls are
varied: imidazopyridine, imidazopyrimidine, oxazine, thiazine, e.t.c. These aryls are with N or O
or S as a Michael donner to be more fitted to receptor and also accompanied with a NO. donor
moiety to overcome classical NSAIDS complications. Herein, the new anti-inflammatory agents
with a hydroxyimoyl or a C-nitroso group (10-15) are prepared. Ulcer liability and

3
histopathological examination of the funds stomach of rats ingested with the prepared
compounds were operated and compared with the references celecoxib and indomethacin. Also
isoxazoles 17 and isoxazolopyridazines 18 were prepared and explored towards their (NO.)
donor activity. Moreover, the research discussed the way of how to 2-aminopyridine and
enaminones reacted with the synthesized hydroximoyl chloride 10 and the structure of the
formed rings are elucidated by 2D NMR. Synthetic protocols were described in the following
Schemes (1-3).
[

4
SO2NH2
CH3
SO2CH3
O
O
O
F3C
N
N
N
SO2NH2
Valdecoxib(2)
celecoxib (1)
Rofecoxib (3)
O
n-Bu O
SO2NH2
N
N
O
N
O
CH3
O
O
R1
ONO2
O
R
NO
O
F3C
N
N
N
N
SO2NH2
N
F3C
SO2NH2
NMI-1093 - (6)
(5)
(4)
R=R1=C1-C6 alkyl or C6-C20 aryl
N
ON
different nitrogen
hetroaryls
as Micheal donners
nitric oxide moiety to
overcome NSIADS side effects
X
pyrazole as a commen feature
of selective COX-2 inhibitors
N
X= N,O,S
N
I
NIASIDS-NO drug hybride
Fig.1: Selective COX-2 ( 1-3), COX-2-NO drug hybrid (4-6) and designed compounds (I).
19
The reaction of 2-Bromo-1-(5-methyl-1-phenyl-1H-pyrazol-4-yl)ethan-1-one (7)
with
dimethylsulfide in refluxing ethanol has afforded dimethyl(2-(5-methyl-1-phenyl-1H-pyrazol-4-
yl)-2-oxoethyl)sulfonium bromide (8). Furthermore, the nitrosation of (8) in dioxane-water
solution acidified with hydrochloric acid gave (Z)-N-hydroxy-2-(5-methyl-1-phenyl-1H-pyrazol-
4-yl)-2-oxoacetimidoyl chloride (10) (Scheme 1). The structure of 10 was confirmed based on

5
elemental analysis, spectral data, and subsequent chemical transformations. That the IR spectrum
showed a broad peak corresponding to (OH) group at 3224 cm-1 in addition to a (conjugated
1
CO) at 1655 cm-1. Also H NMR showed a signelt D2O exchangeable peak at δ 13.45 due to (
OH). The 13C NMR showed two characteristic peaks one at δ 146.65 due to (C=NOH) and the
other at δ 176.98 due to (C=O).
O
N
Br
O
N
Br
S(CH3)2
Cl
N
a
O
N
b
O
N
- (CH3)2S
OH
N
N
c
N
CH3
N
7
10
8
9
Scheme 1: synthesis of (Z)-2-chloro-2-(hydroxyimino)-1-(5-methyl-1-phenyl-1H-pyrazol-4-
yl)ethanone (4); reagents and conditions: (a) Br2/CHCl3, 2hr;(b) (CH3) S, EtOH, 2hr;(c)
2
NaNO2 / HCl, Dioxane-H2O.
Treatment of 10 with 2-aminopyridine in ethanol afforded one isolated product according to TLC
and could be formulated as drawn (11A-C). Structure 11C was ruled out based on the IR
spectrum, which showed no peaks between region 1650-1750 cm-1 due to absence of CO group.
Also, structure 11B was discarded due to 2D NMR that showed NOE correlation between methyl
proton and pyridine H-5. Pyridine-H-5 which appear as a doublet peak at δ 9.87 as with J = 8.4
was close in space enough at a distance of (2.61, 2.73, 4.08 À) with the methyl proton those
appeared as singlet at δ 2.84 and their correlation appeared in 2D NOESY scan. Therefore
postulate structure 11A
named as
3-(5-methyl-1-phenyl-1H-pyrazol-4-yl)-2-
nitrosoimidazo[1,2-a]pyridine (Fig. 2). Analogously, compound 10 was reacted with 2-

6
aminopyrimidine
to
give
3-(5-methyl-1-phenyl-1H-pyrazol-4-yl)-2-nitrosoimidazo[1,2-
a]pyrimidine (12) (Scheme 2).
N
O
Cl
O
ON
N
O N
N
N
N
O
H2N
N OH
O
a
N
N
N
N
N
N
N
-HCl
A
11
10
b
N
N
O
ON
N
H2N
N
N
N
O
N
NO
N
N
N
N
N
11 (B)
N
12
Scheme 2: synthesis of imidazopyridine 11 and imidazopyrimidine 12. Reagents and
conditions: (a) 2-aminopyridine, ethanol, room temperature.; (b) 2-aminopyrimidine,
ethanol, room temperature

7
NOESY scan expansion of 11
Fig.2: 3D representation and NOESY scan expansion of compound 11.

8
1
Structure 12 was elucidated via elemental analysis and spectral data. Thus H NMR showed
three signals, one at δ 7.24-7.28 as triplet due to imidazopyrimidine H-6, and two doublet at δ
8.91, 10.09 for imidazopyrimidine H-7 and imidazopyrimidine H-5. And the 13C NMR
(DEPTQ) showed additional peaks due to imidazopyrimidine ring at: δ
114.47
(imidazopyrimidine C-6), 133.84 (imidazopyrimidne C-5), 143.46 (imidazopyrimidne C-5),
149.20 (imidazopyrimidine C-3), 151.04 (imidazopyrimidine C-2), 158.03 (imidazopyrimidine
C-8a) and 158.43 (imidazopyrimidine C-7) respectively. Also, Compound 10 was reacted with
each of 2-aminophenol , 2-aminothiophenol , 3-hydroxy-2-aminopyridine or o-phenylenediamine
in ethanol afforded 3-(5-methyl-1-phenyl-1H-pyrazol-4-yl)-2-nitroso-4H-benzo[b][1,4]oxazine
(13a), 3-(5-methyl-1-phenyl-1H-pyrazol-4-yl)-2-nitroso-4H-benzo[b][1,4]thiazine (13b), 3-(5-
methyl-1-phenyl-1H-pyrazol-4-yl)-2-nitroso-4H-pyrido[3,2-b][1,4]oxazine (13c) 2-(5-methyl-1-
phenyl-1H-pyrazol-4-yl)-3-nitroso-1,4-dihydroquinoxaline (13d), respectively (Scheme 3).

9
O
X = CH,Y = O
N
NH
O
N
13a
N
O
N
N
NH
O
O
Cl
NH2
YH
X = N, Y = O
N
+
N
13b
N OH
X
N
N
S
N
NH
O
10
X = CH,Y = S
13c
N
N
N
N
HN
N
O
N
X = CH,Y = NH
O
NH
O
N
N
N
N
13d
14
Scheme 3: synthesis of C-nitroso pyrazoles 13a-d and 14; reagents and conditions: (a)
ethanol; (b)H2O2/acetic acid.
Compound 14 formation was excluded due to the disappearance of any (CO) at region 1650-
1750 cm-1 (scheme 3). Structures of 13a-d were confirmed based on elemental analyses and
spectral data. On the other hand, 2-(5-methyl-1-phenyl-1H-pyrazol-4-yl) -3-nitroso-1,4-
dihydroquinoxaline (13d) easily converted to 2-(5-methyl-1-phenyl-1H-pyrazol-4-yl) -3-
nitrosoquinoxaline (15) via oxidation with hydrogen oxide in acetic acid. Finally, hydroximoyl

10
chloride 10 was reacted with the appropriate 3-(dimethylamino)-1-phenylprop-2-en-1-one (16a),
3-(dimethylamino)-1-(p-tolyl)prop-2-en-1-one (16b) and 3-(dimethylamino)-1-(5-methyl-1-
20
phenyl-4,5-dihydro-1H-pyrazol-4-yl)prop-2-en-1-one (16c)
afforded (4-benzoylisoxazol-3-
yl)(5-methyl-1-phenyl-1H-pyrazol-4-yl)methanone (17a), (5-methyl-1-phenyl-1H-pyrazol-4-
yl)(4-(4-methylbenzoyl)isoxazol-3-yl)methanone (17b) and isoxazole-3,4-diylbis((5-methyl-1-
phenyl-1H-pyrazol-4-yl)methanone) (17c), respectively (Scheme 4).
R
O
O
O
N
N
N
16
Cl
O
N OH
+
R
a
O
RCOCH=CHN(CH )
N
3 2
N
O
15
O
R
N
N
N
N
10
N
b
O
N
N
N
17a-c
18a-c
15-19,a, R = phenyl
b, R = 4-(CH3)-C6H4-
c,R= 1-(5-methyl-1-phenyl-1H-pyrazol-4-yl)ethanone
Scheme 4: synthesis of isoxazoles 17a-c and pyradiazinoisoxazoles 18a-c; reagents and
conditions: (a)toluene, triethylamine, stirr,12hr; (b): NH2NH2,ethanol, reflux 2hr.

11
Structures of 17a-c were elucidated by elemental analysis, spectral data (IR, 1H NMR, 2D NMR
and MS) and chemical transportation. From 2D NMR, structure 18 is the only form due to
isoxazolo H-5 correlate with one carbonyl that appeared at δc 186.29 of (C=O) and obligate the
presence of form 17 (Fig. 3). Also structure 16 was out because of further cyclization of the
diketoform with hydrazine hydrate. Thus, treatment of compounds 17a-c with hydrazine hydrate
in ethanol gave
7-(5-methyl-1-phenyl-1H-pyrazol-4-yl)-4-phenylisoxazolo[3,4-d]pyridazine
(18a), 7-(5-methyl-1-phenyl-1H-pyrazol-4-yl)-4-(p-tolyl)isoxazolo[3,4-d]pyridazine (18b), 4,7-
bis(5-methyl-1-phenyl-1H-pyrazol-4-yl)isoxazolo[3,4-d]pyridazine (18c), respectively.
R
R
O
H
O
O
O
O
O
N
N
CH3
17a in form A
CH3
N
N
N
N
17 a in form B
Ph
Ph
HMBC scan of 17a in form A
Fig 3: compound 17a possible forms and HMBC extension scan

12
The in vitro cyclooxygenases inhibition assays were proceeded to explore the effectiveness of
21
synthesized pyrazoles and the ratio of COX -1/2 was calculated . A colorimetric enzyme
immune assay kit (EIA) was used. The doses cause 50% inhibition for both COX-1 and COX-2
were determined and listed in Table 1. The selectivity index (SI) that defined as the ratio of
COX-1/COX-2 was determined that through it, we could expect the safety of the synthesized
compounds. Regarding COX-1 inhibition, most synthesized pyrazoles showed lower potency to
cyclooxygenase-1 that would introduce drugs with good safety profiles. The data obtained
compared to the reference drug celecoxib (1). Generally, all synthesized pyrazoles showed
weakly potent activity against COX-1. 17a, 18b and 18c were with lowest inhibition activity
with (IC50 = 11.1, 10.17, 10.11 µM respectively) and all belonged to isoxazolo group only or
fused with pyridazine. Six pyrazoles (10, 11, 13a, 13c, 13d, and 15) showed IC50 with the range
(7.72-9.80 µM) which is more favorable than celecoxib (IC50 = 7.70). These pyrazoles belonged
to the C - nitroso group. While 12, 13b, 17b and 18a were with lower potency against COX-1.
Regarding COX-2 inhibition activity, 17b and 18a were the most reactive compounds with IC50
= 1.12, 0.78 µM in sequent which are more appreciable than celecoxib (IC50 = 1.14 µM). Also,
these active pyrazoles is belonged to isoxazolo or isoxazolpyridazine groups. Selectivity index
calculated and it showed that 17a, 18a and 18b were more selective than celecoxib.The anti-
inflammatory activity (AI) of new designed NO donors (10, 11, 12, 13a-d, 15, 17 a, b and 18a,
b) were evaluated using carragenan rat paw edema assay method 22. Doses 25, 50, 100 mg/kg of
all synthesized compounds and celecoxib as a standard were administrated orally 1hr before
carragenan injection. The paw thickness was measured 3hr after drug administration's for each
dose. The % percent inhibition in the rat paw edema model was measured and the dose causing

13
50% inhibition (ED50) were calculated for each of the tested compounds and the reference
celecoxib. The data obtained were listed in Table 1.
Table 1: In vitro COX-1 and COX-2 inhibition, and anti-inflammatory (AI) data for
synthesized pyrazoles and the reference drug celecoxib (1).
SIb
COX-1/COX-2
Paw thickness change
Mean SEM
(%Inhibition of paw edema) at
dose 50mg/kg (AI)c
ED50
(mg/kg)
Compound
no
COX-1
IC50 (µM)a
COX-2
IC50
(µM)a
2.86 0.19
(0%)
(50%)
control
10
11
-
-
-
-
9.80
9.20
6.80
8.60
5.60
7.72
9.22
9.20
11.1
4.52
9.11
5.88
10.17
10.11
7.70
2.33
1.75
1.55
4.20
1.22
1.83
1.61
6.60
1.49
1.12
4.50
0.78
1.28
6.88
1.14
4.20
5.25
4.38
2.04
4.59
4.21
5.77
1.39
7.44
4.03
2.02
7.84
7.94
1.46
6.75
1.33 0.24***
50
35
38
45
35
45
40
40
61
56
ND
50
46
ND
35
0.93 0.24*** (65%)
1.02 0.16*** (62%)
1.17 0.17*** (56%)
0.97 0.12*** (64%)
12
13a
13b
13c
13d
15
17a
17b
17c
18a
18b
1.18 0.11***
(56%)
1.15 0.16 *** (57%)
1.10 0.06*** (59%)
1.58 0.04***
1.47 0.15***
NDd
(41%)
(45%)
1.33 0.24***
1.23 0.12***
ND
(50%)
(54%)
18c
Celecoxib
(1)
0.75 0.09 *** (65%)
a The in vitro test compound concentration required to produce 50% inhibition of COX-1 or
COX-2. The result (IC50, µM) is the mean of two determinations acquired using an ovine COX-
1/ COX-2 assay Kit (Cayman Chemicals Inc., Ann Arbor, MI, USA) and the deviation from the
mean is <10% of the mean value.
b In Vitro COX-2 selectivity index (COX-1 IC50/COX-2 IC50).
cInhibitory activity in a carrageenan-induced rat paw edema assay. The results are expressed as
the ED50 value (mg/Kg) at 2 hours after oral administration of the test compound.
d ND = not determined.
Values represent mean SEM (n=3). Significance level ***P<0.001as compared to the control
group.

14
Pyrazoles 11 and 13b were equally effective with celecoxib and showed (ED50= 35 mg/kg) as
that of celecoxib and both belong to C-nitroso group. Pyrazoles 10, 12, 13a, 13b, 13d, 15 and
18b showed moderate in vivo inhibitory activity with an ED50 range (38-46 mg/kg). Isoxazolo
pyrazoles 17a, 17b and 18a were the least potent pyrazoles with ED50 = 61, 56 and 50 mg/kg
respectively.
The nitric oxide generation of all prepared compounds after 2hr administration of 50mg/kg for
three successive days was measured as total (NO2/NO3) in blood serum of rats. A colorimetric
assay determination Griees reagent 23. Most of synthesized pyrazoles showed a significant
increase in serum NO. level (Table 2). Regarding C-nitroso series, they were the most effective
compounds. Pyrazoles 11 and 17 showed 3 fold increase in serum nitric oxide levels.
Hydroxyimoyl chloride 10, 12, 17a and 17d showed two fold increase and this through the direct
liberation of nitric oxide at blood PH= 7.4. For isoxazolo and isoxazolopyridazine groups,
24
literature survey stated that isoxazolo moiety is a proper functional group for a nitric oxide
donation but nothing about isoxazolopyridazine moiety. Isoxazolo compounds 17a-c showed a
variable nitric oxide release. The highest one is 17a with a benzoyl moiety (serum NOxlevel
372.00 10.95 = µmol/l) and the activity remain when methylbenzoyl moiety is added (serum
NOx level 18b, 346.50 52.43). But when substituted with a bulk group pyrazole moiety (17c)
the activity decline to two third of that of 17a. Isoxazolopyridazine group 18a-c showed
moderate activity when compared to the control and they the least active series of all prepared
compounds (Table 2) this through the direct liberation of nitric oxide as a nitroxyl which the
reduced formof NO in blood at PH= 7.4 according to the following mechanism (Fig.4).

15
Table 2: serum NOxlevel, ulcer index and Relative ulcerogenicity to celecoxib of
synthesized diaryl pyazoles (10-18).
Compound
no
Serum NOxlevel
(µmol/l)
Ulcer index
(UI)
Relative
ulcerogenicity to
celecoxib (RU)
147.90 31.76
315.50 19.35*
402.60 23.03***
386.30 24.02**
335.30 13.53*
415.30 24.63***
348.30 32.05*
370.10 44.51**
140.10 88.11
372.00 10.95**
346.50 52.43*
286.20 47.85
314.40 29.69
273.10 20.34
318.8 53.20*
0.00
5.00*
2. 00
0.33
2.00
0.33
0.33
2.33
3.00
0.33
1.33
0.33
1.76
0.33
1.33
0.33
0.00
15.15
6.06
1
6.06
1
Control
10
11
12
13a
13b
13c
13d
15
17a
17b
17c
18a
18b
18c
1
7.06
9.09
1
4.03
1
5.06
1
4.03
1
Celecoxib
-
For nitric oxide determination: values represent mean SEM (n=3). Significance level
*P<0.05, **P<0.01, ***P<0.001 as compared to the control group.
For ulcer liability experiment values represent mean (n=3). Significance level *P< 0.05 as
compared to the control group.
O
O
O
PH= 7.4
-HCl
R
Cl
decomposition
HNO
R
1-
NO
R
H2O
OH
acylnitroso
NOH
hydroxyimoyl chloride
PH= 7.4
H2O
ArOH
HNO
ArC-nitroso
2-
Fig.4: Liberation of nitric oxide at blood PH= 7.4 mechanism.

16
25
Ulcer liability
of was operated after oral administration of 50mg/kg of all prepared
diarylpyrazoles and the reference celecoxib (Table 2). Both ulcer index (UI) and relative
ulcerogenicity (RU) with that of celecoxib were also calculated. Most of prepared compounds
showed lower reactivity as ulcer inducers. 12, 13b and 13c all belonged to a C - nitroso group
substituted with pyrimidine, benzoxazine and benzothiazine moiety and they are equivalent to
celecoxib (UI= 0.33; RU = 1). In addition to 17a (isoxazole substituted with a benzoyl), 17c
(isoxazole with a pyrazole) and 18b (isoxazolopyridazine) showed similar reactivity as
celecoxib. The hydroximoyl chloride 10 is with the highest ulcer liability (UI = 5; RU = 15.15)
this could be related to a spontaneous liberation of HCl after oral administration so increase the
incidence of ulcer occurring. The rest of compounds showed moderate reactivity with the range
of UI (1.33-2.33) and RU (4.03-9.09). pyrazoles 12, 13b, 13c, 17c and 18b were safe as
celecoxib with UI = 1.
Stomach sections were taken for the control rats and that treated with the prepared compounds
and the two references a gastric safe (celecoxib) and the ulcer inducer (indomethacin). These
slides were stained with hematoxylin and eosins stains 26. The prepared slides were examined
microscopically and some picture were picked (Fig.5). The control (Fig.5A) showed a normal
appearance with no lesion anywhere. Indomethacin (Fig.5B) showed a marked loss of mucous
and occasionally noticed abdominal perforations in some places. For pyrazoles 12 (Fig.5E) and
18c (Fig.5F) and celecoxib (Fig.5C) showed no characteristic lesions, but a minimum
vasocapillary dilation was noticed rarely in few slides, for hydroximoyl chloride 10 (Fig.5D) a
characteristic loss of mucosal layer and vasocapillary dilation were seen in certain sites and this
could be due to the direct action of HCl that is spontaneously released after compound intake.

17
Most of examining compounds showed a more safety profile than that of indomethacin and
equally to celecoxib as compounds 12, 13b, 13c, 17c and 18b.
B
C
A
D
E
F
Fig 5: photomicrograph of fundic stomach mucosa : (A) for control, (B) for indomethacin,
(C) for celecoxib, (D) for 10 , (E) for 13c and (F) for 12.
Molecular docking simulation experiments were performed for the most active compounds 12,
13b and 18b towards COX-2 receptor. These compounds showed IC50 1.55, 1.22 and 0.78 µM
as data given in Table 1. The docking experiment was done to investigate the binding reaction of
the compounds with the COX-2 receptor site. Molecular Operating Environment (MOE, 2010,
Version 8, Chemical Computing Group Inc., Montreal, Quebec, Canada) was the software of
choice for molecular docking study. The X-ray coordinates were deposited in the RCSB Protein
Data Bank with codes (PDB: ID 3LN1) 27. The Co-crystal active ligand celecoxib bound at

18
COX-2 isoform. That legend form three hydrogen bonding at N: of (–SO2NH2) COX-2 isoform
at three amino acids (Ser 339, Leu338 and Gln 177) in a distance equal to 2.77,1.83 and 2.03 A˚
respectively (Fig.6, Table 3):. The energy score for receptor active ligand was found to be -
13.86Kcal/mol. The docking of the most active 18b, as a co-active ligand to COX-2 isoform
showed only one hydrogen bond of donor type at pyrazole: N2 with the amino acid Tyr 341at a
distance of (3.05 A˚) that could explain its lower in vivo anti-inflammatory activity (54%
inhibition paw edema) that of celecoxib (65% inhibition paw edema). Also 18b showed higher
COX selectivity (7.94) than celecoxib (6.75) but its lower in vivo reactivity could related to its
lower oral absorption. 18b showed a lower energy score (-11.77 Kcal/mol) but it had a different
way of hydrogen bonding differ from Upon examining 13a, showed three hydrogen bonding of
pyrazole N2, N of (N=O) and oxygen (N=O) with amino acids (Arg 106, Tyr 341, Arg 499) at a
distance of (2.84, 2.59, 2.40 A˚in sequent), as a hypothesis, the higher no of hydrogen bonding
of compound 13a explain its higher in vivo anti-inflammatory (64%) the same as celecoxib .
Finally the imidazopyrimdine 12 showed two hydrogen bonds of (:N=O) and oxygen (N=O:)
with the amino acid (Arg 106) at a distance (2.28, 2.49 A˚) (Fig 7) and an energy score -12.40
Kcal/mol. 12 in vivo result (62%) encourage the pervious stated hypothesis. Although of lower
energy score and higher in vivo reactivity of both 12 and 13a (62, 65% inhibition paw edema),
they showed lower COX selectivity (4.38 , 2.03 in sequent) and this could be related their good
ADME profiles. From the experiment result, induction of different heteroaryl with Micheal
donor moiety induced the compound –receptor interaction through HB of donor type.

19
A
B
Fig6: Binding of Celecoxib bound inside COX-2. (A) 2D interaction of the proposed
binding mode of the active site of COX-2 , the most important amino acids are shown
together with their respective numbers. It forms threeb H-bonds with Ser 339, Leu338 and
Gln 177; (B) 3D interaction of celecoxib.

20
A
B
Fig 7: Binding of 12 bound inside COX-2. (A) 2D interaction of the proposed binding mode
of the active site of COX-2, the most important amino acid is shown together with its
respective numbers. It forms two H-bonds with Arg 106; (B) 3D interaction of 12.

21
Table 3: docking outputs (E-scores and hydrogen bonding ) of compounds 12,13b and 18b.
Compound no.
E-score
No. of
Hydrogen
Distance(A˚)
Hydrogen bonding residues/
bonds
2
bond type
Arg 106H.don
12
-12.40
2.28
Arg 106H.don,
Arg 106 H.don.
2.49
2.84
13b
-13.22
3
Tyr 341 H.don,
2.59
Arg 499 H.don
Tyr 341 H.don.
2.40
3.05
18b
-11.77
-13.86
1
3
Celecoxib
Gln 178H.don,
Leu 338H.don,
Ser 339H.don
2.77
1.83
2.03
As a conclusion C-nitroso pyrazoles 11-14 prove their excellent nitric oxide donor activity.
isoxazolopyrazoles 17a-d and isoxazolopyridazine 18a-d also increased serum nitrite /nitrate
level but with a low significance. Hydroximoyl chloride (10) is the most ulcerogenic
compound. In vitro COX1\2 inhibition activity was evaluated and proved promising activity. For
COX-2 inhibition, compounds 17b and 18a were the most reactive compounds with IC50= 1.12,
0.78 µM in sequent that more reactive than celecoxib itself. In vivo activities as anti-
inflammatory Pyrazoles 11 and 13a were equally effective with celecoxib. Most the compounds
were safe as ulcer inducer and most of the compounds showed excellent nitric oxide (NO.) level
except isoxazolopyridazine. Regio-isomeric reaction of 2-aminopyridine and enaminone with
the hydroximoyl chloride 10 were investigated and proved by 2D NMR. Drug receptor
interaction was investigated and their mechanism was also illustrated. All expermintal details are
in references and notes part. 28-35

22
Acknowledgements
The authors are grateful to all members of the Pharmaceutical Organic Chemistry and the
NMR committee , Faculty of pharmacy, Beni-suef university for all supports during proceeding
of research.
References and notes
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266.
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Graneto M.J. ; Lee, L.F. ; Malecha, J.W.; Miyashiro J.M. , J. Med. Chem. 1997,40, 1347.
3. Dongie, J. M. ; Suprano C.T. ; D. Pratico , J. Med. Chem. 2007, 48, 2251.
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Fukuto, J. M. , Pharmacology & therapeutics 2007, 113 (2), 442.
5. Gao, Y-J ; Lu,C. ; Su, L-Y; Sharma, A. M. ; Lee, R. M. K. W., British Journal of
Pharmacology 2007, 151 (3), 323.
6. Wallace, J. L. ; Ignarro, L. J.; Fiorucci, S. , Nat. Rev. Drug Disc. 2002, 1(5), 375-382.
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8. Stamler, J.S.; Chapel H. , NC (US patent) NO.7.030, 238B2 Apr.18, 2006.
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chemistry 2017, 17 (8), 941.
10. Dahwan,V. ; Schwalb, D. J. ; Shumway, M. J. ; Warren, M. C.; Wexler, R. S.; Zemtesva,
I. S. ; Zifcak, B. M. ; Janero, D. R., Free Radical Biology & Medicine 2005, 39, 1191.
11. Brandi, A. ; Cordero, F. M. ; De Sarlo, F. ; Gandolfi, R. ; Rastelli A. ; Bagatti, M.,

23
Tetrahedron 1992,48, 3323.
12. Kanemasa, S. ; Kobayashi, S. ; Nishiuchi, M. ; Yamamoto H. ; Wada, E. Tetrahedron
letters 1991, 32 , 6367.
13. Kandeel, M.M.; Kamal, A.M. ; Abdelall, E. K. A. ; Elshemy, H. A. H. Eur. J. Med.
Chem. 2013, 59, 183.
14. Kandeel, M. M. ; Ali, S.M.; Abdelall, E.K.A.; Abdelgawad M.A. ; Lamie, P.F. Der
Pharma Chemica, 2012, 4, 1704.
15. Kandeel, M.M. ; Abdelhameid, M.K. ; Abdelall E. K. A; Labib, M.B. Chemical and
Pharmaceutical Bulletin 2013, 61(6) 637.
16. Abdellatif K. R. A.; Abdelall, E. K. A. ; Fadaly, W. A. A.; Kamel G. M., Bioorg. Med.
Chem. Lett. 2016, 26, 406.
17. Elgemeie, G. H.; El-Enany, M.M. ; Ismail M.M. ; Abdelall, E. K. A. Nucleosides,
Nucleotides and Nucleic Acids 2002, 21 (6-7) 477.
18. Salem, E.M. ; Darweesh, A.F; Mekky, A. E. M. ; Farag, A.M. ; Elwahy, A.H.M. J.
Heterocycl.Chem. 2016, 00, 00. DOI: 10.1002/jhet. 257
19. Shaaban, M.R.; Eldebss, T.M.A. ; Darweeh A.F. ; Farag, A.M., J. Heterocycl. Chem,
2008, 45, 1739.
20. Abdelall E. K. A. ; Kamel, G.M. Eur. J. Med. Chem. 2016 , 118 , 250.
21. Elshemy , H. A.H. ; Abdelall , E. K.A.; Azouz; A.A.; Moawad, A.; Ali, W. A.M.; Safwat,
N. M., Eur. J. Med. Chem. 2017, 127, 10.
22. Abdelall, E.K.A. ; Lamie P.F. ; Ali, W.A.M., Bioorg. Med. Chem. Lett. 2016, 26, 2893.
23. Miranda, K. M. ; Espey M. G. ; Wink, D. A. Nitric oxide 2001, 5 (1), 62-71.
24. Ghidini, E. ; Capelli, Carnini, A. M. ; C.; Cenacchi, V. ; Marchini, G. ; Virdis, A. ;

24
Italia, A. ; Facchinetti, F., Steroids 2015 ,95, 88-95.
25. Cho, C. H. ; Ogle, C. W. , European journal of pharmacology 1979, 55 (1), 23.
26. Bancroft, J. D. ; Gamble, M., Theory and practice of histological techniques. 2008, 6th
edition, North Hollywood, USA.
27. http: //www.rcsb.org/pdb/explore/explore.do?structureId=1CX2
28. Synthesis of (Z) -2-chloro-2-(hydroxyimino)-1-(5-methyl-1-phenyl-1H-pyrazol-4-
yl)ethanone (10)
To a mixture of sodium nitrite (3.5gm, 50 mmol) and compound 8 (13.6g, 40mmol) in dioxane /
water (1:1), Conc HCl (50 mmol) is added at 250 and stirred for 3 h. A pale yellow solid was
formed which separated by filtration and crystallized from CHCl3/CH3OH (1:1).
0
Yellow shiny solid, Yield 66%; mp: 227-229 C; IR (KBr, cm-1) 3224 (OH), 3059 (CH), 2833
(CH), 1655 (CO). 1H NMR (CDCl3): δ 2.65 (s, 3H, CH3), 7.45-7.55 (m, 1H, Phenyl H-4), 7.57-
7.62( m, 4H, phenyl H-2, H-6, H-3 and H-5), 8.70 (s,1H, pyrazole H-3), 13.45 (s, OH, D2O
exchangeable); 13C NMR (DEPTQ): δ 12.69 (CH3), 117.57 (pyrazole C-4), 125.60 (phenylC-
2&C-6), 129.39( phenyl C-4) , 129.72 (phenyl C-3& C-5), 137.41 (phenyl C-1), 139.14
(pyrazole C-5),143.72 (pyrazole C-3), 146.65 (C=NOH), 176.98 (C=O). MS (m/z, %): 263.05
[(M)+., 1.66%], 185[100%]: Anal. Calcd. For C12H10ClN3O2: C, 54.66; H, 3.82; N, 15.94, Found:
C, 54.39; H, 3.95; N, 16.09%.
General method for synthesis of compounds 11 and 12
A mixture of 10 (1.3 gm, 5 mmol) and 2-aminopyridine or 2-aminopyrimidine (5 mmol for each)
in ethanol (30 mL) is stirred for 3 h. The resulting solid was filtered and crystallized from
ethanol / ethylacetate (1:1).
2-(5-Methyl-1-phenyl-1H-pyrazol-4-yl)-3-nitrosoimidazo[1,2-a]pyridine (11)

25
Green cubes, Yield 66%; MP: 193-195 0C; IR (KBr, cm-1): 3224 (OH), 3059 (CH), 2833 (CH),
1
1382 (NO). H NMR (DMSO): δ 2.84 (s, 3H, CH3), 7.47 (dd, 1H, imidazopyridine H-6), 7.52-
7.56 (m, 1H, phenyl-4), 7.62-7.67 (m, 4H, Phenyl H-2, H-6, H-3 and H-5), 7.97-8.07 (m, 2H,
imidazopyridine H-7 and H-8), 8.56 (s, 1H, pyrazole H-3), 9.87 (d J = 8.4, imidazopyridine H-
13
5). C NMR (DEPTQ): δ 13.21 (CH3), 112.88 (pyrazole C-4), 117.44 (imidazopyridine C-8),
120.41( imidazopyridine C-6), 125.96 (phenylC-2&C-6), 126.76 (imidazopyridine C-7), 129.07
(phenyl C-4), 129.86 (phenyl C-3& C-5), 138.38 (imidazopyridine C-5), 139.07 (pyrazole C-5),
142.19 (phenyl C-1), 142.48 (pyrazole C-3), 147.01(imidazopyridine C-2), 153.25
(imidazopyridine C-3),156.31 (imidazopyridine C-9). Anal. Calcd. For C17H13N5O C, 67.32; H,
4.32; N, 23.09, Found: C, 67.01; H, 4.03; N, 22.98%. HMBC and HSQC and NOESY in
supplementary materials.
2-(5-Methyl-1-phenyl-1H-pyrazol-4-yl)-3-nitrosoimidazo[1,2-a]pyrimidine (12)
Green cubes, Yield 56%; mp: 180-182 0C; IR (KBr, cm-1) 3055 (CH), 2833 (CH), 1382 (NO).1H
NMR (CDCl3): δ 2.95(s, 3H, CH3), 7.24-7.28 (m, 1H imidazopyrimidine H-6), 7.46-7.51 (m, 1H,
Phenyl H-4), 7.52-7.57 (m, 4H, Phenyl H-2, H-6, H-3 and H-5), 8.84 (s, 1H, pyrazole H-3), 8.91
(d , 1H, imidazopyrimidine H-7), 10.09 (d, 1H, imidazopyrimidine H-5);13C NMR (DEPTQ): δ
13.50 (CH3), 112.62 (pyrazole C-4), 114.47 (imidazopyrimidine C-6), 125.59 (phenyl C-2&C-6),
128.79 (phenylC-4), 129.35(phenyl C-3&C-5), 133.84 (imidazopyrimidne C-5), 138.79
(phenylC-1) 143.42 (pyrazole C-3),143.46 (imidazopyrimidne C-5) 149.20 (imidazopyrimidine
C-3), 151.04 (imidazopyrimidine C-2), 158.03 (imidazopyrimidine C-8a), 158.43
(imidazopyrimidine C-7); MS(m/z, %): 304.22 [(M)+., 2.55%], 77 [100%]: Anal. Calcd. For
C16H12N6O; C, 63.15; H, 3.97; N, 27.62, Found: C, 62.99; H, 3.88; N, 27.98%.
General procedure for synthesis of 13a-d

26
A mixture of 5 (1.3 gm, 5 mmol) and the appropriate of 2-aminophenol, 2-amino-3-
hydroxypyridine , 2-aminothiophenol or o-phenylenediamine (5 mmol) in ethanol / dioxane (20
mL, 1:1) is stirred for 2 h. The resulting solid was filtered and crystallized from ethanol / ethyl
acetate (1:1).
3-(5-Methyl-1-phenyl-1H-pyrazol-4-yl)-2-nitroso-4H-benzo[b][1,4]oxazine (13a).
Buff needles, Yield 66%; mp: 158-160 0C; IR(KBr, cm-1) 3059 (CH), 2868 (CH), 1350 (NO). 1H
NMR (DMSO): δ 2.65 (s, 3H, CH3), 7.45-7.55 (m, 2H, Phenyl H-4 and benzoxazine H-5), 7.57-
7.65 (m, 6H, phenyl H-2, H-6, H-3, H-5 ,benzoxazine H-7 and pyrazole-H-3), 7.94 (d J= 8, 1H,
benzoxazine H-8), 8.05 (d, J = 8, 1H, benzoxazine H-6), 8.90 (s, 1H, NH); 13C NMR (DEPTQ):
δ 12.96 (CH3), 112.44 (benzoxazine C-5), 118.44 (pyrazole C-4), 122.49 (benzoxazine C-8),
125.98 (phenyl C-2&C-6), 126.37 (benzoxazine C-7), 128.88 (benzoxazine C-6), 128.88
(phenyl-4), 129.55 (phenyl C-3& C-5), 138.35 (phenyl C-1), 140.59 (benzoxazine C-2), 144.13
(pyrazole C-5), 146.46 (pyrazole C-3), 150.32 (benzoxazine C-3), 158.14 (benzoxazine C-4a),
174.27 (benzoxazine C-8a). MS (m/z, %): 318.27 [(M)+., 0.87%], 185[100%]: Anal. Calcd. For
C18H14N4O2: C, 67.91; H, 4.43; N, 17.60, Found: C, 64.09; H, 4.22; N, 17.92 %.
3-(5-Methyl-1-phenyl-1H-pyrazol-4-yl)-2-nitroso-4H-pyrido[3,2-b][1,4]oxazine (13b).
Red needles, Yield 66%; mp: 254-256 0C; IR(KBr, cm-1) 3065 (CH), 2878 (CH), 1340 (NO).
1H NMR (DMSO): δ 2.86 (s, 3H, CH3), 7.26-7.29 (m, 1H, pyridoxazine H-7), 7.38(d J= 8HZ,
1H, pyridoxazine H-8), 7.51-7.55 (m, 1H, Phenyl H-4 ), 7.59-7.68 (m, 4H, phenyl H-2, H-6, H-3
, H-5 ), 8.52 (s,1H, pyrazole H-3), 9.46 (d J = 8 HZ,1H, pyridoxazine H-6); 11.29 (NH, D2O
exchangeable); 13C NMR (DEPTQ): δ 13.13 (CH3),
113.14 (pyrazole C-4), 118.05
(pyridoxazine C-7) 120.38 (pyridoxazine C-8), 121.20 (pyridoxazine C-6), 125.76 (phenyl C-
2&C-6), 128.96 (phenylC-4), 129.83 (phenyl C-3& C-5), 139.22 (phenyl C-1), 140.24

27
(pyridoxazine C-3), 141.92 (pyrazole C-5), 142.28 (pyrazole C-3), 146.55 (pyridoxazine C-2),
153.87 (pyridoxazine C-8a), 174.27 (pyridoxazine C-4a); Anal. Calcd. For C17H13N5O2: C,
63.94; H, 4.10; N, 21.93, Found: C, 63.77; H, 4.33; N, 21.79 %.
3-(5-Methyl-1-phenyl-1H-pyrazol-4-yl)-2-nitroso-4H-benzo[b][1,4]thiazine (13c)
Yellow needles, Yield 76%; mp: 279-2810C; IR (KBr, cm-1) 3062 (CH), 2870 (CH), 1355 (NO).
1H NMR (DMSO): δ 2.57 (s, 3H, CH3),7.30-7.38(m, 2H, benzothiazine H-5&H-7), 7.45-7.47
(m, 2H, benzothiazine H-6&H-8), 7.50-7.55(m, 1H, Phenyl H-4 ), 7.57-7.89 (m, 4H, phenyl H-2,
H-6, H-3, H-5), 8.16 (s, 1H, pyrazole H-3) 13C NMR (DEPTQ): δ 13.24 (CH3), 117.59 (pyrazole
C-4), 122.05 (benzthiazine C-2), 125.82 (phenyl C-2&C-6), 126.30 (benzothiazine C-5), 127.64
(benzothiazine C-7), 128.47 (benzothiazine C-6), 128.73 (phenylC-4), 129.75 (phenyl C-3& C-
5), 131.01 (benzothiazine C-8), 138.50 (phenyl C-1), 139.43 (benzothiazine C-3), 140.91
(benzothiazine 8a), 141.09 (pyrazole C-5), 142.50 (pyrazole C-3), 147.72 (benzothiazine C-4a).
Anal. Calcd. For C18H14N4OS: C, 64.65; H, 4.22; N, 16.75, Found: C, 64.30; H, 4.01; N, 16.62
%.
1,4-Dihydro-2-(5-methyl-1-phenyl-1H-pyrazol-4-yl)-3-nitrosoquinoxaline (13d)
0
Yellow solid, Yield 76%; mp: 230-232 C; IR (KBr, cm-1) 3321 (NH), 3059 (CH), 2868 (CH),
1350 (NO). 1H NMR (DMSO): δ 2.61 (s, 3H, CH3), 6.96 (m, 1H, quinoxaline H-5), 7.20 (m, 1H,
quinoxaline H-8), 7.32 (m, 1H, quinoxaline H-6), 7.41(m, 1H, quinoxaline H-7), 7.57-7.62 (m,
5H, phenyl H-4, H-2, H-6, H-3, H-5 ), 8.38 (s, 1H, pyrazole H-3), 10.30 (s, 1H, NH, D2O
exchangeable), 10.55 (S, 1H, NH, D2O exchangeable); 13C NMR (DEPTQ): δ 13.32 (CH3),
112.44 (quinoxaline C-5), 116.42 (pyrazole C-4), 121.44 (quinoxaline C-8), 125.79 (phenyl C-2
& C-6), 127.92 (quinoxaline C-6), 128.71 (quinoxaline C-7), 129.02 (phenyl C-4), 129.76
(phenyl C-3& C-5),132.15 (quinoxaline C-2), 132.34 (quinoxaline C-3), 139.37 (phenyl C-1),

28
140.66 (quinoxaline C-4a), 141.85 (pyrazole C-5), 142.44 (pyrazole C-3), 149.24 (quinoxaline
C-8a). MS (m/z, %): 317.34 [(M)+., 41.23%], 300.20 [100%]: Anal. Calcd. For C18H15N5O: C,
68.13; H, 4.76; N, 22.07, Found: C, 68.02; H, 4.62; N, 22.22 %.
Synthesis of 2-(5-Methyl-1-phenyl-1H-pyrazol-4-yl)-3-nitrosoquinoxaline (15)
A solution of 16d (0.5 gm) in 10 mL of acetic acid and hydrogen peroxide (30%, 5 mL) is added
and stirred overnight. The reaction mixture was poured in water and the solid precipitate was
filtered and crystallized from DMF/ethanol (1:1).
Brown solid, Yield 46%; mp: 270-2720C; IR(KBr, cm-1) 3059 (CH), 2868 (CH), 1350 (NO). 1H
NMR (DMSO): δ 2.63 (s, 3H, CH3), 7.48-7.53 (m, 5H, phenyl H-4, H-2, H-6, H-3, H-5 ),7.59-
7.60 (t, 1H, quinoxaline H-5), 7.99-8.01 (t, 1H, quinoxaline H-8), 8.06 (d J = 7.2 Hz, 1H,
quinoxaline H-6), 8.21 (d J = 7.2 Hz, 1H, quinoxaline H-7), 8.30 (s, 1H, pyrazole H-3). MS
(m/z, %): 314.32 [(M-1)+., 13.37%], 316.22[(M+1)+., 2.15%], 233.27 [100%]: Anal. Calcd. For
C18H13N5O: C, 68.56; H, 4.16; N, 22.21, Found: C, 68.02; H, 4.62; N, 22.42 %.
General procedure for synthesis of 17a-c.
A mixture of 10 (1.3 gm, 5 mmol), appropriate enaminone 16a-c (5 mmol) and triethylamine
(0.5 gm, 0.75 mL) in toluene (30 mL) and stirred for 6 h at room temperature. The reaction
mixture is evaporated and the oil residue was dissolved in ethyl acetate and crystallized from
hexane / ethyl acetate mixture (1:1).
1-(5-Methyl-1-phenyl-1H-pyrazol-4-carbonyl)-isoxazolo-4yl-(phenyl)methanone (17a)
0
Creamy white solid, Yield 56%; MP: 205-207 C; IR (KBr, cm-1) 3059 (CH), 2868 (CH), 1682
1
(CO), 1624 (C=N), 1600 (C=C). H NMR (DMSO): δ 2.55 (s, 3H, CH3), 7.59-7.60 (m, 7H,
phenyl H-4, H-2, H-6, H-3, H-5 and benzoyl H-3&H-5), 7.71-7.75 (m, 1H, benzoyl H-4), 7.95
(d, J = 7.6, benzoyl H-2&H-6), 8.10 (s, 1H, pyrazole H-3), 9.80 (s, 1H, isoxazole H-5); 13C NMR

29
(DEPTQ): δ 12.66 (CH3), 119.94 (pyrazole C-4 & isoxazole C-4), 125.30 (phenyl C-2 & C-6),
129.30 (phenyl C-4), 129.47 (benzoyl C-3 & C-5), 129.56 (phenyl C-3 & C-5), 129.63 (benzoyl
C-2 & C-6), 129.88 (benzoyl C-4), 134.43 (phenyl C-1), 137.17 (benzoylC-1), 138.28 (pyrazole
C-5), 145.32 (pyrazole C-3), 160.88 (isoxazole C-3), 164.02 (isoxazole C-5), 179.66 (C=O),
186.29 (C=O). HMBC&HSQC are in supplementary materials. MS (m/z, %): 357.19 [(M)+.,
19.85%], 185.12 [100%]: Anal. Calcd. For C21H15N3O3: C, 70.58; H, 4.23; N, 11.76, Found: C,
70.32; H, 4.66; N, 11.32%).
1-(5-Methyl-1-phenyl-1H-pyrazol-4-carbonyl)-isoxazolo-4yl-(4-methylphenyl)methanone (17b).
0
Creamy solid, Yield 56%; MP: 190-192 C; IR (KBr, cm-1) 3072 (CH), 2885 (CH), 1680 (CO),
1622 (C=N), 1605 (C=C). 1H NMR (DMSO): δ 2.42 (s, 3H, CH3), 2.55 (s, 3H, CH3), 7.39 (d, J =
8Hz, 2H, 4-methylbenzoyl H-2 & H-6), 7.55-7.61 (m, 5H, phenyl H-4, H-3, H-5,H-2&H-6 ),
8.12 (d, J = 8Hz, 2H, 4-methylbenzoyl H-2 & H-6), 8.16 (s,1H, pyrazole H-3), 9.76 (s, 1H,
isoxazole H-5); 13C NMR (DEPTQ): δ 12.66 (CH3), 18.99(CH3),119.92 (pyrazole C-4), 120.02
(isoxaole C-4), 125.91 (phenyl C-2&C-6), 129.58 (phenyl C-4), 129.80 (4-methyl benzoylC-
3&C-5), 129.90 (4-methylbenzoylC-4, phenylC-3&C-5), 130.04 (4-methyl benzoylC-2&C-6),
134.62 (phenyl C-1), 138.25 (4-methyl benzoylC-1), 143.78 (pyrazole C-3), 145.14 (pyrazole C-
5), 160.89 (isoxazole C-3), 163.66 (isoxazole C-5), 179.69 (C=O), 185.79(C=O). Anal. Calcd.
For C22H17N3O3: C, 71.15; H, 4.61; N, 11.31, Found: C, 71.55; H, 4.86; N, 11.61%).
1-(5-Methyl-1-phenyl-1H-pyrazol-4-carbonyl)-isoxazolo-4yl-(1-(5-methyl-1-phenyl-1H-
pyrazol-4-yl)methanone (17c)
0
Creamy solid, Yield 56%; MP: 174-176 C; IR (KBr, cm-1) 3063 (CH), 2865 (CH), 1675 (CO),
1
1626 (C=N), 1605 (C=C). H NMR (DMSO): δ 2.53 (s, 3H, CH3), 2.60 (s, 3H, CH3), 7.54-7.62
(m, 10H, 2X( phenyl H-4, H-2, H-6, H-3, H-5 ), 8.05 (s, 1H, pyrazole H-3), 8.36 (s,1H,
pyrazoloyl H-3), 9.90 (s, 1H, isoxazol-H-5); 13C NMR (DEPTQ): δ 12.32 (CH3), 13.84 (CH3),