249
CHAPTER V
SYNTHESIS, CHACTERISATION AND
BIOLOGICAL ACTIVITY OF NOVEL
PYRAZOLINE DERIVATIVES.
250
INTRODUCTION
PYRAZOLES
Pyrazole refers both to the class of simple aromatic ring organic
compounds of the heterocyclic series characterized by a 5-
memberedring structure composed of three carbon atoms and two
nitrogen atoms in adjacent positions and to the unsubstituted
parent compound. Being so composed and having pharmacological
effects on humans, they are classified as alkaloids although they are
not known to occur in nature.
Pyrazoles are produced synthetically through the reaction of α,
β-unsaturated aldehydes with hydrazine and subsequent
dehydrogenation. Hydrogenation is a chemical reaction in which
unsaturated bonds between carbon atoms are reduced by
attachment of a hydrogen atom to each carbon.
NN
H
NH2NH
2O
It may also be prepared by the union of diazomethane with
acetylene and by warming the acetal of propargyl aldehyde with an
aqueous solution of hydrazine sulphate. It crystallizes as colourless
needles, which is very stable and behaves as a weak base. It does
not combine with the alkyl iodides. On oxidation with potassium
permanganate the C-alkyl-derivatives give carboxylic acids, whilst
251
the N-phenyl derivatives frequently split off the phenyl group
(especially if it be amidated) and have it replaced by hydrogen. On
reduction, the pyrazoles with a free NH group are scarcely affected,
whilst the N-phenyl derivatives give pyrazolines, or by the use of very
strong reducing agents the ring is ruptured and
trimethylenediamine derivatives are formed. They yield substitution
derivatives with the halogens, bromine being the most effective. The
chloro-derivatives are most readily prepared from the pyrazolones by
the action of phosphorus oxychloride.
The pyrazole carboxylic acids may also be obtained by
condensing 1, 3-diketone or oxymethylene ketone carboxylic esters
with hydrazines, or the diazo fatty esters with acetylene
dicarboxylic esters. The dihydro pyrazoles or pyrazolines are less
stable than the pyrazoles and are more like unsaturated
compounds. They are weak bases, which are only soluble in
concentrated acids. On reduction, they yield pyrazolidines, or the
ring is broken; and when oxidized they form blue or red colouring
matters. The carboxylic acids show a remarkable behavior on
heating, the nitrogen is entirely eliminated, and trimethylene
carboxylic acids are obtained. Pyrazoline is a colourless liquid,
which boils at 144 °C. It may be prepared by the action of
diazomethane on ethylene.
252
The pyrazolones (ketodihydropyrazoles) first prepared from the
elimination of the elements of alcohol from the hydrazones of 0-
ketonic acids; or on the oxidation of the pyrazolidinones with ferric
chloride. Three types are possible with the formulae: H2C CO I; NH
HC: N Pyrazol-5-one. They form salts with both acids and bases,
and yield benzylidine and isonitroso derivatives. Pyrazolone is
obtained by the condensation of hydrazine with formyl acetic ester.
It is a colourless crystalline solid, which melts at 164 °C. 1-Phenyl-
3-methyl pyrazole-5-one, is antipyrine. The isomeric 1-phenyl-5-
methylpyrazol-3-one is formed by condensing acetoacetic ester
with acetophenyl hydrazine in the presence of phosphorus oxy
chloride, or by the action of ferric chloride on the corresponding
pyrazolidone, which is produced by condensing phenyl hydrazine
with an O-halogen butyric acid. When methylated it yields
isoantipyrine, an isomer of antipyrine, which is more poisonous.
Pyrazolidines are tetrahydro pyrazoles. The N-phenyl
derivative, from sodium phenyl hydrazine and trimethylene
bromide, is oil, which readily oxidizes to phenyl pyrazoline on
exposure. Isomeric compounds may arise here when phenyl
hydrazine is used, the keto-group taking either the 3 or 5 position;
thus with ß-iodopropionic acid 1-phenylpyrazolidine-5-one is
formed, whilst potassium 1,3-iodopropionate gives the 3-
compound. Isomers of this type may be distinguished by the fact
253
that the pyrazolidine-5-one compounds are basic, whilst the 3-
compounds are acidic. The simplest member of the series,
pyrazolidine-5-one, is a liquid that is formed by the action of
hydrazine on acrylic acid. The 3.5-pyrazolidones are the cyclic
hydrazides of the malonic acid series.
Thiopyrazoles have been obtained by the action of an
aqueous or alcoholic solution of the methyl chloride or iodide of
phenyl methyl chloro pyrazole on a solution of an alkaline
hydrosulphide into which carbon bisulphide has been passed; or
by the action of sodium thiosulphate on antipyrine hydrochloride
or a similar compound1.
Pyrazoles are used for their analgesic, anti-inflammatory,
antipyretic, antiarrhythmic, tranquilizing, muscle relaxing,
psychoanaleptic, anticonvulsant, monoamineoxidase inhibiting,
antidiabetic, antimicrobial2 and antibacterial activities.
Structurally related compounds are pyrazoline and pyrazolidine.
NN
NN
H
NN
H
H
The above three represent heterocyclic nomenclature to
pyrazolines require that nitrogen atoms to be numbered one and
two in each structure. Substituted 1-pyrazolines are numbered to
produce the lower locations, or in the case of complicated
254
structures are produce the simplest name consistent with clarity of
meaning. Numbering of the 2-pyrazolins begins with the amino
nitrogen and pyrazolines are numbered to obtain for the double
bond the lower of the two possible numbers. Thus, here this
structure will be referred as:
NN
1
2
34
5
The pyrazoles, pyrazolines and the pyrazolidines form an
interesting class of compounds showing diverse biological
activities. The pyrazoline and pyrazolidines can be considered as
the hydrogenated, compounds of pyrazoles. Many pyrazoles and
pyrazolines surveyed in the literature have been screened against
various microorganism and their pharmacological activities also
compared.
PYRAZOLINES: CHEMISTRY AND METHOD OF SYNTHESIS
There are different methods that have been reported for the
synthesis of pyrazolines.
Rao et al3 synthesized pyrazolines through cycloaddition of
diazomethane to chalcone, which in turn were obtained by crossed
aldol condensation of appropriately substituted benzaldehyde and
acetophenones. Earlier reporters4,5 has suggested two alternative
structures (2) and (3) for these pyrazolines.
255
R
RHC=CH-C
R
RO
R
R
NHN
C
O R
R
R
R
NN
C
O R
R
(1)
(2)(3)
1
2
3
1
1
2
2
3
3
Mustafa et al6 were report that 5-arylcryloyl-6-hydroxy-4, 7-
dimethoxy-benzofuran react with hydrazine hydrate in ethanol to
give hydrazones which on boiling with acetic acid yield the
corresponding pyrazolines.
Aziz et al7 reported the reaction between hydrazine hydrate
and 5-aryl, acryloyl-6-methoxy-benzofuran was shown to yield the
corresponding 2-pyrazoline via., 1.4 addition shown to yield the
corresponding 2-pyrazoline via., 1.2 addition in acetic acid, the
isomerization of 1, 4 addition product to 1, 2 addition product in
boiling acetic acid was also reported by these workers.
Wadodkar et al8,9 by taking account of above synthesis is
reported that 2-hydroxy chalcone (1) reacts with hydrazine
hydride, but the products were identified as 5-aryl-3-(2-hydroxy
256
phenyl)-2-pyrazoline(2).The formation of pyrazoline is possible only
by 1,2 addition.
On boiling in acetic acid this pyrazoline did not undergo
isomerization as reported by Musfate et al10 5-aryl-1-acetyl-3- (2-
hydroxy phenyl) 2-pyrazoline (3).
It was been also reported that hydroxyl chalcone (4) with
hydrazine hydrate in dimethyl formamide or hyrazone
hydrochloride and aqueous sodium acetate or sodium hydroxide in
ethanol were similar to (5) and not the corresponding isomer of (6).
Further it has been also reported that hydroxyl chalcone failed to
react with hydrazine hydrochloride in ethanol alone, even on
prolonged refluxing (6 to 8 h).
OHR
RC-CH=CH
O
RNNO
HbHa
HxR
R
H
RNNOH
Hb
Ha
HxR
R
Ac
R
3 3
3
1
2
1
2
1
2
4 5 6
Hydroxyl chalcone reacts with hydrazine hydrochloride in dimethyl
formamide (soft base) to give (7), which remains unchanged on prolonged boiling
in acetic acid.
C-CH=CH
OHR
R
O
NNOHR
R Hb
HaNH2NH
2.HCl
DM
1
2 2
1
7
257
Osman et al11 has reported that substituted cinnamolyl
chloride (8) reacts with 2-aryl-5H-oxazolo, (4,5b) phenoxazoline (9)
in dioxane in the presence of triethylamine to give (10). 5-
substituted benzalacetyl oxazolo phenoxazine derivative (10) react
easily with hydrazine hydride and Phenyl hydrazine to give
corresponding pyrazolines.
It has been reported that interaction of compound (10) with
hydrazides under suitable conditions gave a variety of pyrazolines.
Hydrazine hydrate itself interacts with (10) in dioxane giving
unstable pyrazolines (11). But when this was treated with glacial
acetic acid, then stable monoacetyl pyrazoline was obtained (12).
258
CH=CHCOCl
N
O
H
N
OAr
N
O
COCH=CH
N
OAr
NH2NH
2H
2O
GLACIAL ACETIC ACID
R
N
O
N
OAr
N N
CO
CH3
N
O
N
OAr
N NC
6H
5
RR
+
9
10
11 12
8
Phenyl hydrazine reacts with (10) in the presence of base
catalyst. The reaction was carried out in diaxone in presence of
piperidine giving N phenyl pyrazoline (12).
Sachchar et al12 synthesized fluorinated heteroaryl
pyrazolines, in this the fluorinated hetero aryl chalcone reacts with
phenyl hydrazine to yield phenyl hydrazone, which immediately
rearranges to the corresponding isomeric, 1-phenyl-3- (substituted
fluro phenyl)-5-heteroaryl pyrazolines(13) in presence of glacial
acetic acid. In this case it was reported that mixture was refluxed
259
in an oil bath at 1200-130oc for 3 to 4 hours. After cooling the
resultant mass was diluted with water and the solid obtained was
recrystallized from ethanol. These derivatives when placed on filter
paper and exposed to bromine vapour tuned green and have
identified as pyrazolines.
CH3COOH
C6H
5NH
2NH
2NN
R
CH=CH-C R
RO
R
120-130o
+
13
1
1
260
Review of Literature
PYRAZOLES:
Dadiboyena et al13 synthesized pyrazole derivatives (14) and
reported their anti bacterial and anti inflammatory activity.
Porter et al14 synthesized Tetrahydroisoquinoline amide (15)
substituted phenyl pyrazoles as selective Bcl-2 inhibitors.
O
N
NH2
NN
Cl
O
N
15
Wahab et al15 synthesized and evaluated antimicrobial
activity of 1-(benzofuran-2-yl)-4-nitro-3-aryl butan-1-ones and 3-
(benzofuran-2-yl)-4,5-dihydro-5-aryl-1- 4-(aryl)-1,3-thiazol-2-yl]-
1H-pyrazoles(16), (17) and (18).
NN
R
NH
CH3
14
261
N NO
N
S
Ar
N NO
NH2
S
OAr
NO2
16 17 18
Prakash et al16 synthesized and evaluated antibacterial
activity of some new 2,3-dimethoxy-3-hydroxy-2- (1-phenyl-3-aryl-
4-pyrazolyl) chromanones(19).
.
O
OOH
OMe
OMeN
N
Ar
O
OOH
Ph
N
N
Ar
Ph
IBD
MeOH
19
Gupta et al17 Synthesized N-aryl-5-amino-4-cyanopyrazole
derivatives (20) as potent xanthine oxidase inhibitors.
R
NN NH
2
CN
HC(OEt)3,Ac
2O
NH3
R
NN
N
N
NH2
20
Kralj et al18 reported a simple synthesis of 4-(2-aminoethyl)-
5-hydroxy-1H-pyrazole (21).
262
N O N O
NMe2
NN
NH2
OH
COPh COPh
21
Pyrazoline as antimicrobial and anti bacterial:
Sachchar et al19 synthesized several 1-phenyl-3-(substituted
fluro phenyl)-5-heteroayl-2-pyrazolines (22) and noted their anti
microbial as well as anti bacterial actions especially against
bacteri-B anthrasis.
N N
RR
R = furyl, thienyl,pyridylR1= F,Cl,OH.CH3
1
22
Many other authors also reported the synthesis and
antibacterial activity of some new pyrazoline derivatives.20-24
Stirrewiberg et al25 reported antibacterial as well as
insecticidal action of some new pyrazoline derivatives (23).
R
NN
R
C
O
NH R
R
23
1
2
3
263
Desai et al26 synthesized several pyrazoline derivatives of
phenothiazines and reported their antibacterial activity and
tuberculostatic activity of some compounds.
Ead et al27 reported the synthesis of two novel series of
pyrazoles and 2-pyrazolines with their antibacterial activity.
Pyrazolines as antifungal:
Sachchar et al28 extended their studies on several pyrazoline
derivatives and reported them as antifungal. Aspergillus niger and
Helmithosporium sativum were employed as fungi for the testing of
fungicidal activity.
Ritcha et al29 synthesized 3, 5 dimethyl -4- nitro pyrazole
and 1, 3, 5 trimethyl-4-nitrosopyrazole compounds and have
reported them as anti fungal agents. Their fungicidal activity
increased by increasing the size of hydrocarbon side group was
also reported by them.
Mitra et al30 reported that 5-pyrazolone and its derivatives 4-
acetyl-2-pyrazoline-5-one and 2-pyrazoline-5-thione (24) were
associated with significant fungicidal activity against the rice born
pathogen Pyricularia oryzae and brown leaf spot pathogen
Helminthosporium oryzae31-33
264
NN O
CO S
N
R
S
24
Mitra et al34 have synthesized 4-N (aryl) amino methyl-2-
pyrazoline-5-one by Mannich reaction of 2-pyrazoline-5-one
derivative. The same authors also reported cobalt II complex with
2-pyrazoline-5-one derivative as having anti fungal activity.
Sadasiva et al35 synthesized several hydroxyl aryl-pyrazole
(25) and tested them for antibacterial as well as anti-fungal activity
where none of the compounds were found to possess anti-bacterial
activity. But all compounds were found to possess anti-fungal
activity. Antifungal activity has been assessed by employing
Drechslera prostrate (Drechs) and Alternatia alternate (Keissler).
All compounds tested could inhibit the spore germination at 30-
mcg/ml upto a maximum level.
N N
OH
R
R
1
25
R=alkyl or aryl R1=H or phenyl
265
Nayak et al36 synthesized several 4,4‟-bis-5-pyrazoline and 4,
4‟,unsaturated products and found their fungicidal activity against
rice blast pathogen Pyricularia oryazae and brown leaf spot
pathogen, Helmithosporium oryzae.
Tiwari et al37 synthesized several 1-acetyl/aroyl-3-methyl-4-
substituted anilido -5-aryl pyrazolines and 3-methyl-4-substituted
anilido-5-aryl pyrazolines and 3-methyl-4-substituted anilido-5-
aryl isoxazolines, and tested against Cephalosporium sacchari,
Helmithosporium oryzne, and saprolegina parasitica, acellya Orion
all the compounds showned remarkable activity.
Mohanthy et al38 reported that 1-phenyl-5-aryl-1-2-
pyrazoline 3-4-thiazolidine-2-one derivative exhibited antifungal
properties.
The fungicidal activity of the compounds was determined by
poisoned food technique at various concentrations. It was also
reported that all compounds inhibit the growth of the fungus
Aspergillus flavours.
Pyrazoline as antiviral:
Sachachar et al39 extended their study on several phenyl1-3-
(substituted fluro phenyl)-5-hetero aryl-2-pyrazoline derivative and
found antiviral activity against Sunn hemp rosette virus (SRV).
266
Pyrazoline as antiserotonin and anti-oedima:
Another derivative Dipyrone was prepared which showed a
strong antiserotonin and anti-oedima activities in rats40 but found
to be clinically disappointing in the treatment of rheumatic
arthritis41. Other compounds like 4-(N-Nicotinyl amino) derevative
was also found to be less toxic and clinically effective in various
inflammation diseases42
Frangnly et al43 have reported anti-inflammatory activity of
some new pyrazoles pyrazolines and 4(3H)-quinazolines.
Derivative of 3, 5-pyrazolindinedione as anti inflammatory:
After the discovery of the 5-pyrazoline derivative as anti-
inflammatory agents, modifications were made on the basic
structure and this resulted in the synthesis of a new compound,
phenyl butazone, which was found to be a potent inhibitor of
inflammation.
A metabolic product of phenyl butazone was found to be
less toxic and equally potent as the parent compound. The result
was the synthesis of oxyphenbutazone.
Modification of the parent compound to increases the acidity
of the molecule resulted in the synthesis of a new derivative sulfin
267
pyrazole44 it was enhanced uricosuric activity and is potent against
gout.
N
N
R
O
O
R
C6H
5
1
2
26
R1 R2
Phenyl butazone -C6H5 -C4H9
Oxyphen butazone -C6H4-OH (P) -C4H9
Sulfin pyrazone -C6H5 -CH2-CH2-S-C6H5
Pyrazole derivative as anti-inflammatory agent:
Coli et al45 reported that pyrazole derivative itself has anti-
inflammatory activity (27) e.g. Benzylame (Tantum)
NN
CH2-Ph
OCH2R
27
R = -CH2-CH2-N (CH3)2, -COCH3
268
Sarangam et al46 have synthesized number of derivative of
pyrazole –(3,4-d) pyrimidine-4-6 diones (28) and screened them for
C.N.S. depression properties and anti-inflammatory activity. It was
also reported that some derivatives showed anti-inflammatory
properties equivalent to aspirin.47
H
N
N
O
ON
N
R
R
1
28
R1= Phenyl, o-Tolyl R2 = Phenyl, o-Tolyl, o-Anisyl
Pyrazole derivatives as antidiabetic:
Froesch et al48 has reported antidiabetic activity in the 5-
methyl pyrazole-3-carbaxilic acid (29).
CH3
NN
COOH
H
29
Pyrazole derivative as vasodilator:
Brunner et al49 have reported vasodilator activity in pyrazole
derivative (30).
269
N
NN
N
CH3
30
Pyrazole derivative as hypoglycemic and hypotensive agent:
Smith et al50 reported that 3, 5-dimethyl pyrazole and 3-
methyl pyrazole-5-carboxilic acid exhibited hypoglycemic activity
(31).
Arya et al51 synthesized several pyrazolidine derivatives and
reported the hypotensive activity. (32)
NN
CH3
CH3
H
NN
CH3
H
HOOC
31
NN
H
C
O
CH2
CH2
NN CH3
CH3
32
270
Pyrazolidone derivative as inhibitor:
Sweeny et al52 reported that pyrazofurin which is a natural
antibiotic was very effective in inhibiting pyrimidine biosynthesis.
Chasin et al53 discovered a potent new compound pyrazole
pyridine derivative (33) that was 60 times more potent as an
inhibitor of rat brain PDE than theophyllin.
N
C2H
5OOC
NH-N=C-(CH3)
NN
33
Novinson et al54 reported a series of pyrazole pyrimidine
derivatives (34,35) and reported them to be inhibitor of PDE from
rabbit lung and beef heart.
NN
NR
R
NNN
N
R
R
EtOOC
H
1
34 35
1
Many compounds containing the pyrazoline nucleus have
been found to show different biological activities.55-58
271
METHODOLOGY
NH2
CH3
CH3
CH3
ONNH
2NH
2.H
2O
NH2
ON
C
O
R
NH2
ON
CH2CO
C2H
5OH
C2H
5OH
CH2
C O
O
C
CH2CONHNH
2
CH CH
N
N
R
+
R1
R
PZ1-14
reflux
Scheme IV
R1
CLH
CLE
CH1-14
24hrs
272
Preparation of Caprolactam hydrazide (CLH):
A mixture of t-Butyl (s)-3-amino benzocaprolactum (CLE,2.9gm,
0.01 mol) and 99% of hydrazine hydrate (0.05 mol) in dry ethanol
(50 ml) was refluxed for 24 Hrs. The solution upon cooling gave a
solid mass that was filtered and recrystallized from chloroform.
(Yield) 58%, MP: 170 0C.
Synthesis of chalcones (CH1-14):
A solution of acetophenone (12gm, 0.1mol) and benzaldehyde
(11gm, 0.1mol) were prepared in 100ml of ethanol separately. The
acetophenone solution was then added to benzaldehyde solution.
The mixture was stirred for 5mins. Then a solution potassium
hydroxide (10%) was added dropwisely until turbidity appears.
Then mixture was stirred for 12hrs. The precipitate obtained was
filtered, washed with water and recrystallized from ethanol.
Following the same procedure other chalcones were also prepared.
Synthesis of pyrazolines:
A mixture of benzocaprolactam hydrazide (2.5gm, 0.01 mol)
and chalcone (2.0gms, 0.01mol) was refluxed for 6hrs in dry
ethanol (50ml) in the presence of 1ml of glacial acetic acid. Upon
cooling the precipitate obtained was filtered, washed with ether
and recrystallized from the ethanol. Following the same procedure
remaining compounds of the series were prepared. The physical
chacteristics of compounds are complied in the table no.17.
273
Table No.17: Chacterisation data of pyrazoline derivatives (PZ1-14):
S.No
.
Code Substituents Molecular
Formula
M.P.
(oC)
Yield
(%) R1 R
01. PZ1 H H C26H26O2N4 152 70
02. PZ2 H 4-N-Dimethylamino- C28H31N5O2 128 72
03. PZ3 4-Chloro- H C26H25O2N4Cl 145 68
04. PZ4 4-Chloro- 4-Chloro- C26H24O2N4Cl2 116 64
05. PZ5 H 4-Methoxy- C27H28O3N4 180 70
06. PZ6 4-Bromo- 4-Methoxy- C27H28O3N4Br 195 66
07. PZ7 4-Chloro- 4-N-Dimethylamino- C28H30O2N5Cl 204 68
08. PZ8 H 2-Nitro- C26H25O4N3 166 72
09. PZ9 2-Chloro- 2-Nitro- C26H24O4N3Cl 175 66
10. PZ10 4-Bromo- H C26H25O2N4Br 188 74
11. PZ11 H 2-Chloro- C26H25O2N4Cl 126 79
12. PZ12 4-Chloro- 4-Methoxy- C27H27O3N4Cl 180 75
13. PZ13 H 4-Chloro- C26H25O2N4Cl 168 73
14. PZ14 4-Chloro- 2-Chloro- C26H24O2N4Cl2 90 60
NH2
ON
CH2CO N
N
R1
R
274
SPECTRAL DATA OF SYNTHESIZED COMPOUNDS:
Compound CLH:
(3-Amino-2-oxo-2,3,4,5-tetrahydro-benzo[b]azepin -1-yl)-acetic acid hydrazide
IR (KBr)cm-1 :
3364 and 3308(NH and NH2), 3060(Ar.C-H str.), 2970, 2943, 2881,
2835 {Ali.C-H str. of CH2 groups(CH2,N-CH2)asymmetric and
symmetric}, 1744(ring C=O), 1668(C=O of CONH2), 1602(C=N),
1602, 1491, 1456(C=C ring str.), 1395, 1366(C-H bending).
1H NMR:
δ 9.2(1H,s,NH of CONH), 6.87-7.64(4H,m,Ar-H), 4.74-3.0 (6H,m,2H
of NH2 of NHNH2+2H of ring C-NH2 + 2H of CH2 of N-CH2), 2.5(1H
of C-H of ring CH-NH2;merged with solvent peak), 2.23-
2.07(2H,s,ring CH2), 1.74(2H,s,ring CH2)
NH2
ON
CH2CONHNH
2
275
13C NMR of compound CLH:
N
NH2
O
O
NH
NH2
57
68 1 2
4
3 11
12
9
10
Sl.No Ppm Values Assignment
1 27.866 1
2 38.623 2
3 49.456 3
4 51.062 4
5 122.722 5
6 126.084 6
7 127.299 7
8 128.875 8
9 135.918 9
10 141.513 10
11 167.537 11
12 174.639 12
276
277
278
279
Compound PZ1:
3-Amino-1-[3-(3,5-diphenyl-4,5-dihydro-pyrazol-1-yl)-2-
oxo-propyl]-1,3,4,5-tetrahydro-benzo[b]azepin-2-one
IR (KBr)cm-1 :
3237(weak broad peak of –NH2), the disappearance of peaks at
3363, 3308 for NH and NH2 of CONHNH2 support the formation
pyrazoline. 2937, 2862 (Ali. C-H str. of CH2 groups), 1744(ring C=O
properly resolved). The change in the nature of the broad peak at
1667(C=O of side chain) may be due to fermi resonance,
1600(C=N), 1558, 1491, 1458(C=C ring str.), 1398(C-N), 758(mono
substituted phenyl rings),
1H NMR :
The disappearance of the signals at 9.2 for NH of CONH and 2H of
NH2 of CONHNH2 of the hydrazide confirm the formation of
pyrazoline. Other important of signals of pyrazoline PZ1 are δ 8.0-
7.0(14H,m, Ar-H), 5.0(2H,s,NH2 of CHNH2 of ring),4.6-
4.2(2H,dd,2H of pyrazoline), 4.0-3.2(6H,m,4H of ring 2xCH2 + 2H
of N-CH2). The C-H of -CHNH2 of ring perhaps merged with solvent
peak.
NH2
ON
CH2CO N
N
280
281
282
Compound PZ2:
3-Amino-1-{3-[5-(3-dimethylamino-phenyl)-3-phenyl-4,5-dihydro pyrazol-1-yl]-2-oxo-propyl}-1,3,4,5-tetrahydro-benzo[b]azepin-2-one
IR(KBr)cm-1 :
3398(NH2), The characteristic absorption peaks for NH and NH2 of
CONH2 of hydrazide have disappeared. 2922,2204(Ali. C-H str.of
CH2and CH3 groups), 1740(ring C=O), 1670 (C=O of side chain),
1599(C=N), 1566, 1521, 1490(C=C ring str.), 1444,1361(C-H
bending of CH2 and CH3 groups), 769,815(substituted benzene).
1H NMR:
Disappreance of signal for NH of CONH of hydrazide. δ
7.8-7.2(13H,m,Ar-H), 6.6(2H,dd,NH2), 4.8 and 4.2(2H,d,2H of
pyrazoline), 3.2(2H,s,2H of N-CH2),3.0-2.6(5H,m,2H of ring + 1H of
CHNH2 + 2H of CH2 of pyrazoline), 2.1(6H,s, 6H of (CH3)2-N),
1.8(2H,s,2H of CH2 of ring)
Mass Spectrum: The molecular ion peak at m/z 482
corresponding to the molecular weight of the compound is
obtained. In addition, it also gives M+1 and M+2 peaks. Incedently
the molecular ion peak at m/z 482 it self is a base peak. Other
prominent fragment ion peaks are obtained at m/z
468,397,380,351,308,289,266,252,217,161,144,132,106,91, 78.
NH2
ON
CH2CO
NN
N
CH3
CH3
283
284
285
286
CompoundPZ3 :
NH
2
ON
CH2CO N
N
Cl
3-Amino-1-{3-[3-(3-chloro-phenyl)-5-phenyl-4,5-dihydro-pyrazol
-1-yl]-2-oxo-propyl}-1,3,4,5-tetrahydro-benzo[b]azepin-2-one
IR(KBr)cm-1:
3358(NH2), 3059 (Ar, C-H str.), 2929, 2860 (Ali C-H str of CH2 and
–NCH2 groups). The carbonyl peaks at near 1740 clearly
resolved for ring C=O.1670(C=O of side chain), 1599(C=N), 1529,
1491, 1460(C=C ring str.), 1400 (C-N), 1359, 1300 (C-H bending of
CH2), 833, 760 (substituted phenyl rings), 588(C-Cl).
1H NMR:
The signal for NH of CONH at δ 9 is not observed. δ 8.4-
6.7(13H,m,Ar-H), 4.9(2H,d,2H of NH2), 4.2 and 4.6(2H,dd,2H of
pyrazoline), 4.3(2H,s,2H of N-CH2), 2.8-2.6 and 2.2-1.8(6H,m,6H of
ring (CH2)2 and C-H +1H of pyrazoline) Due to the poor solubility of
compound the signals are not clearly appeared. How ever the
disappearance of characteristic hydrazide peaks in the NMR
spectra of pyrazoline supports the formation of pyrazoline. The
structure is further confirmed by its mass spectrum.
287
Mass:
The molecular ion peak of the compound is observed at m/z 471
corresponding to the molecular weight of the compound. In
addition it gives M+1, M+2 peaks. Indecently this is the base peak.
Further the fact that the peak heights of M+1 and M+2 peaks in
the ratio 3:1 confirm the presence of Cl in the compound. The
compound under investigation contains chlorine atom in its
molecule.
288
289
290
291
Compound PZ4:
NH2
ON
CH2CO
ClN
N
Cl
3-Amino-1-{3-[3,5-bis-(3-chloro-phenyl)-4,5-dihydro-pyrazol
-1-yl]-2-oxo-propyl}-1,3,4,5-tetrahydro-benzo[b]azepin-2-one
IR (KBr)cm-1 : 3358(NH2), 3059 (Ar. C-H str.), 2929, 2860(Ali. C-H
str. of CH2 and –NCH2 groups). The carbonyl peaks at near 1740
clearly resolved for ring C=O. 1670(C=O of side chain), 1599(C=N),
529, 491, 460(C=C ring str.), 1400(C-N), 1359,1300 (C-H bending
of CH2), 833, 760(substituted phenyl rings), 588(C-Cl).
1H NMR: The disappearance of peak for NH of CONH of hydraide
supports the formation of pyrazoline. δ 8.2-7(12H,m, Ar-H),
4.9(2H,d,2H of NH2), 4.4-4.6 and 4.2-4.0(2H,dd,2H of pyrazoline),
4.3(2H,s,2H of N-CH2), 2.8-1.8(6H,m,4H of 2xCH2 + 1H of CH-NH2
+ 1H of pyrazoline).
Mass Spectrum: The molecular ion peak of the compound is
observed at m/z 505 corresponding to the molecular weight of the
compound. This is also a base peak. M+, M+1, M+2 peaks are also
observed. The peak heights of M+1and M+2 peaks are in the ratio
3:1.This suggests presence of Cl and confirms the formation and
structure of the compound.
292
13C NMR of compound PZ-4:
N
NH2
O
NO
N
Cl
Cl
1 2
6
22
4 21
35
20
13
13
11
11 18
14 9
9 12
12
7
19
158
10
a
PZ-4
Sl.No Ppm Values Assignment
1 20.419 1
2 21.641 2
3 27.671 3
4 36.911 4
5 37.79 5
6 50.447 6
7 118.49 7
8 122.784 8
9 127.28 9
10 127.478 10
11 128.732 11
12 128.883 12
13 129.518 13
14 130.455 14
15 130.629 15
16 133.826 16
17 134.676 17
18 135.7.1 18
19 136.075 a
20 137.974 19
21 142.971 20
22 172.463 21
23 187.995 22
293
294
295
296
297
Compound PZ6:
3-Amino-1-{3-[3-(3-bromo-phenyl)-5-(3-methoxy-phenyl)-4,5-dihydro pyrazol-1-yl]-2-oxo-propyl}-1,3,4,5-tetrahydro-benzo[b]azepin-2-one
IR (KBr)cm-1 :
The peak for NH and NH2 of hydrazide are disappeared. The
NH2 of CHNH2 perhaps merged with the broad peak appearing
between 3250 and 3064 (Ar. C-H str.), 2935, 2841(Ali C-H str. CH2
of OCH3 groups), the peak for C=O of ring around 1740 clearly
resolved. The peak at 1567 may be due to C=O of side chain. 1593
(C=N), 1510, 1458(C=C ring str.), 1421(C-N), 1396, 1332(C-H
Bending), 1031 (C-O-C) 819,738(substituted phenyl ring), 665 (C-
Br).
1H NMR:
The disappearance of signal for NH of CONH of hydrazide is
observed. δ 8.1-6.8(12H,m, Ar-H), 5.5(2H,d,2H of NH2),5.2-5.3 and
4.9-4.7(2H,dd,2H of pyrazoline), 4.2(2H,d,2H of N-CH2),
3.8(3H,s,3H of OCH3), 3.7-3.5(1H,m,1H of CH-NH2), 3.2-3.0 and
2.2-2.0(5H,m,4H of 2xCH2 of ring +1H of pyrazoline).
NH2
ON
CH2CO
OMeN
N
Br
298
Mass Spectrum:
The mass spectrum of the compound at m/z 546 corresponding to
the molecular weight of the compound. It is very surprising to see
hear that the peaks at m/z 333, 291 and 259 show 100% intensity
showing the stability of fragment ions as base peak. Other
prominent peaks are observed at m/z 475, 409, 355, 315, 277,
219, 203, 185, 144, 130, 106, 91,78.
299
300
301
302
Compound PZ7 :
NH2
ON
CH2CO
N
CH3
CH3
N
N
Cl
PZ7
3-Amino-1-{3-[3-(3-chloro-phenyl)-5-(3-dimethylamino-phenyl)-4,5-dihydro-
pyrazol-1-yl]-2-oxo-propyl}-1,3,4,5-tetrahydro-benzo[b]azepin-2-one
IR (KBr)cm-1 :
3213(NH2 of ring), peaks for NH, NH2 of hydrazide disappeared.
2935, 2820(Ali C-H str. of CH2 and CH3 groups), 1672 (C=O of side
chain), 1600(C=N), 1523, 1489, 1460(C=C ring str.), 1431(C-N),
1364, 1338(C-H bending of CH2 and CH3 groups), 812,
736(substituted phenyl ring), 523(C-Cl).
1H NMR:
The signal for NH of CONH of hydrazide at down field is
disappeared. δ 6.6-8.2(12H,m, Ar-H), 4.9-1.6(18H,m, 2H of NH2 +
2H of NCH2 + 2H of pyrazoline + 4H of 2 x CH2 of ring + 1H of
CHNH2 + 6H of N(CH3)2 + 1H of pyrazoline).
Mass Spectrum:
The molecular ion peak of the compound is observed at m/z 516
corresponding to the molecular weight of the compound. M+, M+1,
303
M+2 peaks are also observed the peak heights of in the ratio 3:1
indicating the presence of Cl. Other prominent peaks are observed
at m/z 502, 431, 380, 342, 298, 286, 235, 217, 144, 132, 106, 91,
and 77.
304
305
306
307
Compound PZ9 :
3-Amino-1-{3-[3-(2-chloro-phenyl)-5-(2-nitro-phenyl)-4,5-dihydro- pyrazol-1-yl]-2-oxo-propyl}-1,3,4,5-tetrahydro-benzo[b]azepin-2-one
IR (KBr) cm-1 :
3310(NH2), 3065(Ar.C-H str.), 2935,2864(C-H str.ofCH2
groups), 1740,1667(ring C=O and C=O of CH2-C=O), 1601(C=N),
1522,1489,1458(C=C ring str.), 1394(C-N), 1522 &1342(-NO2),
832(1,4-disubstitued phenyl ring), 592(C-Cl)
1H NMR:
δ The signal for NH of CONH (hydrazide) at downfield is
disappeared.6.6-8.8(12H,m,Ar-H), 5.0-4.1(2H,d,2H of NH2), 4.6-4.5
and 4.2-4.1(2H,dd, 2H of pyrazoline), 4.3(2H,s,2H of CH2-N), 2.9-
3.0(1H,m,1H of CH-NH2),2.6(1H,m,1H of pyrazoline), 2.2(2H,d,2H
of ring CH2), 1.8(2H,d,2H of ring CH2).
Mass Spectrum:
The molecular ion peak is observed at m/z 518 corresponding
to the molecular weight of the compound. The M+2 peak at m/z
520 also observed. The peak heights of M+ and M+2 appeared to be
NH2
ON
CH2CO
O2N
N
N
ClPZ9
308
in the ratio of 3:1 due to the presence of chlorine atom in the
molecule. The prominant fragment ion peaks are also observed at
m/z 513, 424, 399,368, 288, 256, 235, 177, 144, 132, 106, 80
etc.,
309
310
311
312
BIOLOGICAL ACTIVITY:
The antibacterial, antifungal and analgesic activities for
Pyrazoline derivatives were carried out by following same protocols
and procedures as mentioned in chapter No.III. The results are
represented in the table no.18, 19 and 20 respectively.
TABLE No.18: Antibacterial activity of synthesized
Pyrazolines (PZ1-14):
*Average triplicate ± Standard deviation Note: ‘-‘denotes no activity, 4-5 mm poor activity, 6-7 mm moderate activity, 8-
9 above good activity.
Sample
Code
*Inhibition zone diameter in mm
B.subtilis B.pumilis E.coli S.aureus
50g 100g 50g 100g 50g 100
g 50g
100
g PZ1 4 6 4 7 4 5 - 5
PZ2 5 8 5 7 4 5 - 5
PZ3 5 7 - 5 4 5 - 6
PZ4 6 8 5 7 4 7 5 8
PZ5 4 6 - 6 4 6 - 5
PZ6 6 8 5 6 4 7 4 5
PZ7 5 7 5 4 4 5 4 4
PZ8 5 7 5 7 4 5 - 5
PZ9 5 8 5 7 4 5 4 6
PZ10 4 5 5 6 5 6 4 4
PZ11 6 7 5 6 5 5 5 5
PZ12 5 8 5 6 4 7 4 7
PZ13 6 7 7 8 - 5 4 5
PZ14 5 7 5 7 4 5 4 6
Ciprofloxacin 6 10 5 9 5 8 6 9
DMF - - - - - - - -
313
Table No.19: Antifungal activity of Pyrazolines (PZ1-14):
Note:- “ – “denote no activity, 06 – 07mm poor activity, 08 – 10mm moderate
activity, 11-12mm good activity.
Sample code
*Inhibition zone diameter in mm
A.Nigier C.Albicans
50g 100g 50g 100g PZ1 05 08 05 08
PZ2 06 10 06 09
PZ3 06 09 06 10
PZ4 07 11 07 11
PZ5 04 07 - 07
PZ6 06 11 05 11
PZ7 07 12 06 11
PZ8 05 10 04 10
PZ9 06 11 06 11
PZ10 05 09 05 09
PZ11 05 09 05 08
PZ12 07 12 - 10
PZ13 05 09 05 08
PZ14 05 11 06 09
Clotrimazole 09 14 08 13
DMF - - - -
314
Table No.20: Analgesic activity of Pyrazolines derivatives:
Treatment
No. of animals
Avg. Wt of Animals (gm)
Avg. Dose (mg)
Basal reaction time (secs.) after
0 min 15 min 30 min
60 min
90 min
120 min
Control gum
acacia 6 34.00 -
3.2 ±0.37
3.81 ±0.473
4.12 ±0.553
4.28 ±0.503
4.87 ±0.324
3.88 ±0.37
2 Standard Pentazoc
in 10 mg/kg
6 28.00 0.28 6.24 ±0.89
12.87 ±1.332
12.91 ±1.320
13.87 ±0.279
13.65 ±0.851
11.83 ±1.04
2
Compound PZ1
6 25.33 2.53 6.35 ±0.35
6.90 ±0.384
8.24 ±0.135
9.52 ±0.453
11.21 ±0.882
9.33 ±0.54
9
Compound PZ4
6 29.33 2.93 5..18
±0.508 8.59
±0.490 9.74
±0.260 12.69
±0.313 10.11
±0.882
9.33 ±0.54
9
Compound PZ5
6 22.00 2.20 6.61
±0.372 7.49
±0.257 8.58
±0.692 8.71
±0.740 9.92
±0.411
8.79 ±0.32
0
Compound PZ7
6 26.00 2.60
6.13
±0.826 8.39
±0.579 9.80
±0.924 11.96 ±1.10
10.584 ±0.493
9.60 ±0.44
8
Compound PZ8
6 20.66 2.06 5.87
±0.775 8.13
±0.288 9.75
±0.316 12.10
±0.919 11.10
±0.684
9.03 ±0.27
7
Compound PZ10
6 27.33 2.73 5.31
±0.374 7.57
±0.487 9.36
±0.553 11.62
±1.146 10.71
±0.666
8.07 ±0.29
8
Compound PZ11
6 27.33 2.73 6.46
±0.743 7.68
±0.349 10.4
±0.794 11.9
±0.971 10.96
±0.346
9.74 ±0.37
0
Compound PZ13
6 29.33 2.93 5.16
±0.632 7.27
±1.505 9.46
±1.368 11.87
±0.078 12.12
±0.675
10.43 ±0.61
9
315
RESULTS AND DISCUSSIONS:
A.Antibacterial activity:
All the pyrazoline derivatives synthesized during the present
investigations were screened for their antibacterial activity against
bacteria B. substils , B.pumilus, E. coil and S. aureus at
concentration 50mg and 100mg by disc diffusion method. The
Ciprofloxacin was used as standard and DMF as solvent control.
Surprisingly all the compounds of the series exhibited excellent
activity almost resembling as that of the standard at the
concentration studied. Hence the detailed toxicity study may be
more beneficial .If the highly active compounds proved to be less
toxic, may be used as topical antibacterial agents in the form of
ointments and gels.
B.Antifungal activity:
All the compounds screened for antibacterial activity were also
screened for antifungal activity against the fungi Aspergillus niger
and C.albicans at the concentration of 50 and 100 µg/ml by
following disc diffusion method. Clotrimazole an antifungal agent
was used as standard. The compounds PZ1. PZ5, PZ8 PZ10, PZ11
PZ13 and PZ14, showed moderate activity, while PZ2, PZ3, PZ4, PZ6,
PZ7 and PZ12, showed significant activity in comparison with
standard against the fungi A. niger at both concentration levels.
Similarly PZ2, PZ3, PZ4, PZ7, PZ9 and PZ14 exhibited good activity
316
in comparison with the standard against C.albicans at both
concentrations. The presence of pyrazoline moiety, substituents
particularly one having N-CO-NH2 in the ring and compact
coplanar structure may be responsible for antifungal activity of
this class of compounds.
C.Analgesic activity:
Pyrazoline series of compounds PZ1, PZ4, PZ5, PZ7, PZ8, PZ10,
PZ11 and PZ13 were screened for their analgesic activity by Eddy‟s
hot plate method. Mice of either sex were used for the study.
Pentazocin 10mg/kg body weight used as standard drug for
comparison of activity. The standard drug showed percentage
analgesic activity as 218.75.All the compounds screened showed
promising analgesic activity. The activity ranges from moderate –
equipotent-highly potent. The compounds PZ4, PZ7, PZ8, and PZ10
exhibit significant activity almost equal to that of the standard. The
compound PZ1, PZ5, PZ7 and PZ11 showed moderate activity.
Surprisingly the compound PZ13 that contains chlorine on the p-
position of aryl ring has shown much superior activity. As the
compounds have shown promising analgesic activity it can be
concluded that the pyrazole moiety still serves as a promising
moiety and may provide more clinically useful compounds, if
molecule is suitably modified. Hence the further study of this
317
series of compounds particularly with reference to the toxicity
studies may result useful compound to the human kind.
318
REFERENCES
1. http://www.nationmaster.com
2. Wahab BFA, Aziz HAA, Ahmed EM. Eur J Med Chem 2008; 44:
2632.
3. Rao BCH, Raju SGV. J Ind Chem Soc 1986; 25: 400.
4. Ellis GP, Thomas IL. J Chem Soc Perkin Trans 1 1973: 2781.
5. Arndt F. Chem Ber 1925; 58: 1612.
6. Mustafa A, Hismat OH. Chem Abstr 1966; 65: 3823.
7. Aziz G, Nosseir MH. Ind J Chem 1976; 15B: 446.
8. Thakare VG, Wadodkar KH. Ind J Chem 1996; 25B: 610.
9. Wadodkar KN. Synthesis in heterocyclic compound, Ph.D,
Thesis Nagpur University, Nagpur,1977.
10. Mustafa A, Fleifel AM. J Org Chem 1959; 24: 1740.
11. Osaman AM, Maghuraby MA. J Ind Chem Soc 1977; 54:
394.
12. Sachchar SP, Singh AK. J Ind Chem Soc 1985; 62: 142.
13. Dadiboyena S, Valente EJ, Hamme AT. Tetrahedron Lett
2009; 50: 291.
14. Porter J, Payne A, Candole B, Ford D, Hutchinson B,
Trevitt G et al. Bioorg Med Chem Lett 2009; 19: 230.
15. Wahab BFA, Aziz HAA, Ahmed EM. Eur J Med Chem 2008;
44: 2632.
319
16. Prakash O, Kumar R, Sehrawat R. Eur J MedChem 2008;
44: 1763.
17. Gupta S, Rodrigues LM, Esteves AP, Campos AMFO,
Nascimento MSJ, Nazareth N et al. Eur J Med Chem 2008;
43: 771.
18. Kralj D, Groselj U, Meden A, Dahmann G, Stanovnik B, Svete
J. Tetrahedron 2007; 63: 11213.
19. Sachchar SP, Singh AK. J Indian Chem Soc 1985; 62: 142.
20. Sharma TC, Bokadia MM. J Indian Chem Soc 1980; 19: 228.
21. Nayak A, Mitra AS. J Indian Chem Soc 1980; 57: 643.
22. Tiwari N, Dwivedi B, Nizamuddin. Bol Chem Farm 1989;
128: 332.
23. Mohanty SK, Sridhar R. J Indian Chem Soc 1977; 15: 1146.
24. Mecalcon SE. Agric Chem 1947; 2: 31.
25. Sirrewiberg WE. Chem Abstr 1978; 89.
26. Desai NC, Trivadi PB, Unnadevi NK, Dave AM, Bhat KN. Ind J
Chem 1993; 32B: 760.
27. Ead HA, Hassaneen HM, Abdallah MA, Mousa HAH. Arch
Pharm Weinheim Ger 1991; 324: 35.
28. Sachchar SP, Singh AK. J Indian Chem Soc 1985; 62: 142.
29. Ritch S, Horsfall JC. Chem Abstr 1952; 56: 11543.
30. Mitra P, Nayak A. J Indian Chem Soc 1982; 59: 1005.
320
31. Burns JJ, Yu TF, Ritterband A, Perel JM, Gutman AB,
Brodie BB. J Pharmacol Exp Ther 1957; 119: 418.
32. Coli B, Silerstrint. Exp Med Therap 1957.
33. Rosmer I, Buliard M. Med Pharmacol Exp 1967; 16: 25.
34. Mitra P, Mitra AG. J Indian Chem Soc 1981; 63: 695.
35. Sadasiva M, Rao R. J Indian Chem Soc 1982; 59: 1104.
36. Nayak A, Mitra AS. J Indian Chem Soc 1980; 57: 643.
37. Tiwari N, Dwivedi B, Nizamuddin. Bol Chem Farm 1989;
128: 332.
38. Mohanty SK, Sridhar R. J Indian Chem Soc 1977; 15: 1146.
39. Sachchar SP, Singh AK. J Indian Chem Soc 1985; 62: 142.
40. Smith LI, Denyes RO. J Am Chem Soc 1936; 58: 304. John
W, Gunther P, Schmeil M. Chem Ber 1938; 71: 2637.
41. Elhabiri M, FigueiredoP, Fougerousse A, Brouillard R.
Tetrahedran Lett 1995; 36: 4611.
42. Eder H, Ansri M. Forsch 1961; 11: 1043.
43. Frangly AM, Chaban I, Khalil MA, Behkit AA. Arch Pharm
Winhim 1990; 325.
44. Burns JJ, Yii. J Pharmac Ep Therap 1957.
45. Coli B, Silerstrint. Exp Med Therap 1957.
46. Saragan S, Somashekara S. J Ind Chem Soc 1976; 53.
47. Jone RG. J Am Chem Soc 1952; 74: 488.
48. Froesch ER, Wolaogel M. Mol Pharmacol 1967; 3: 429.
321
49. Brunner HR, Eichenberger K. Experimenta 1966; 22: 208.
50. Smith DL, Forist AA, Dulin WE. J Med Chem 1965; 8: 350.
51. Arya VP, Grewal RS. Experimenta 1967; 23: 824.
52. Sweeny MJ, Davis FA, Gutowski GE, Hamill RL, Hoffman
DH, Poore GA. Cancer Res 1973; 33: 2619.
53. Chasin M, Harris DN, Phillips MB, Sidney M. Hess Bio Chem
Pharmacol 1972; 21: 2443.
54. Navinson T, Scholten H. Am Chem Soc, Abstr Med 1972;
52.
55. Kerdawy MM, Agemy A. Acta Pol Pharm 1975; 7:105.
56. Wrezicino U. Acta Pol Pharm 1975; 30: 433.
57. Gover RK, Moore JD. Phyto Pathology 1962; 52: 876.
58. Nayak A, DasS. J Indian Chem Soc 1977; 54: 485.