Synthesis and Antioxidant Activity of Some New Binary Pyrazoles Containing Core Phenothiazine Moiety

Article abstract of DOI:10.1002/jhet.2716

α,β-Unsaturated ketones containing phenothiazine moiety 4, 5, and 7 were synthesized by condensation of 2-acetylphenothiazine (1) with different aryl aldehydes 2, 3 and dimethylformamide dimethylaceal. Pyrazoles 11, 18, 20, 22, 23, 24, 25, 26, 28, 29, 31,

Full text of DOI:10.1002/jhet.2716

Month 2016
Synthesis and Antioxidant Activity of Some New Binary Pyrazoles
Containing Core Phenothiazine Moiety
Wafaa S. Hamama, Moustafa A. Gouda, Hadwah A. Kamal El-din,a and Hanafi H. Zooroba
a
a,b
*
aDepartment of Chemistry, Faculty of Science, Mansoura University, El-Gomhoria Street, 35516, Mansoura, Egypt
bDepartment of Chemistry, Faculty of Science and Arts, Ulla, Taibah University, Medina, Kingdom of Saudi Arabia
*
E-mail: wshamama53@gmail.com
Received February 19, 2016
DOI 10.1002/jhet.2716
Published online 00 Month 2016 in Wiley Online Library (wileyonlinelibrary.com).
α,β-Unsaturated ketones containing phenothiazine moiety 4, 5, and 7 were synthesized by condensation of 2-
acetylphenothiazine (1) with different aryl aldehydes 2, 3 and dimethylformamide dimethylaceal. Pyrazoles 11,
18, 20, 22, 23–26, 28, 29, 31, 32 and oxazole 34 skeletons were also synthesized by 1,3-dipolar cycloaddition
reactions of α,β-unsaturated ketones with different nucleophilic reagents. Formylpyrazole derivative 36 was
synthesized through Vilsmeier–Haack reaction of phenylhydrazone 35b. Newly synthesized compounds were
screened for antioxidant activity. The data showed clearly that most of the compounds anchored to phenothiazine
moiety displayed good antioxidant activity using ABTS method. Furthermore, compounds 11, 14, and 28 exhibited
high protection against DNA damage induced by the bleomycin iron complex.
J. Heterocyclic Chem., 00, 00 (2016).
INTRODUCTION
may arise in a biological system after increasing exposure to
oxidants, so the antioxidants play a major role in the protection
of biological systems against threats. Different types of antiox-
idants such as vitamins C and E, glutathione, lipoic acid, and
butylated phenols, were widely used in different fields of
industry and medicine to interrupt radical-chain oxidation
processes that attract a high scientific interest [19,20].
In view of the aforementioned facts and in continuation of
our interest in the synthesis of biologically active heterocyclic
[21–23], we report herein the synthesis and antioxidant
evaluation of some novel pyrazoles incorporating phenothia-
zine moiety through different linkages. This combination
was suggested in an attempt to investigate the influence of
such hybridization and structure variation on the anticipated
biological activities, hoping to reach a more potent antioxidant
agent.
Phenothiazine derivatives are well known for their central
nervous system activity [1]. They occur in various neuroleptic
drugs, for example, chlorpromazine, and antihistaminic drugs,
for example, promethazine [2] (Fig. 1). They describe the
largest of the five main classes of neuroleptic antipsychotic
drugs [2].
Phenothiazines represent a key motif in heterocyclic
chemistry and occupy a prime place in medicinal chemistry
due to their competence to exhibit a wide range of pharmaco-
logical activities such as tranquilizer [3], anti-inflammatory
[4], antimalarial [5], anti-psychotropic [6], antimicrobial [7],
anti-tubercular [8,9], antitumor [10,11], antihistaminic [12],
and analgesic [13,14].
Furthermore, pyrazole ring is present as the core in a variety
of leading drugs such as celebrex [15], viagra [16], or
rimonabant. Moreover, 4-aminoantipyrine is used for the
protection against oxidative stress as well as prophylactic of
some diseases including cancer, and these are important
directions in medicine and biochemistry [17,18].
Reactive oxygen species, including free radicals led to a
decrease in the antioxidant capacity and may generate other
reactive species that damage the living cell. Oxidative stress
RESULTS AND DISCUSSION
Chemistry. The synthetic strategies adopted to obtain the
corresponding phenothiazines are depicted in Schemes 1–6.
α,β-unsaturated ketones (4) and (5) were prepared according
to the similar reported method via Claisen–Schimidt
© 2016 Wiley Periodicals, Inc.

W. S. Hamama, M. A. Gouda, H. A. Kamal El-din, and H. H. Zoorob
Vol 000
intermediate compound 18 towards amines was studied.
Thus, its reaction with hydrazine hydrate (19) in boiling
EtOH/AcOH mixture gave pyrazole derivative 20. Also,
treatment of the cyclohexenone derivative 18 with hydroxyl-
amine hydrochloride (21) in boiling alcohol in the presence
of sodium acetate led to the formation of the corresponding
oxime 22 (Scheme 3).
Figure 1. Phenothiazine derivatives are well known as neuroleptic drugs.
Moreover, condensation reaction of α,β-unsaturated
ketones 5 with hydrazine hydrate (19) in ethanol afforded
the corresponding bispyrazole derivative 23. Although
reaction of 5 with phenylhydrazine (8) or hydrazine
hydrate (19) in AcOH respectively, afforded the
correspoding bis pyrazoles derivative 24 and 25. While,
refluxing of 23 in AcOH/Ac2O gave the bisacetylated de-
rivative 26 (Scheme 4).
In a similar manner, compound 4or 5 reacted with
thiosemicarbazide (27) to give the thioamides 28 and 29
which up on refluxed with phenacyl bromide (30) in
ethanol gave the corresponding binary thiazoles 31 and
32, respectively (Scheme 5). Furthermore, the isoxazole
derivative 34 was achieved through reaction of 5 with
aqueous solution of hydroxylamine hydrochloride (21)
and sodium acetate (Scheme 5).
condensation of compound 1 with benzaldehyde (2), or 1,3-
diphenyl-1H-pyrazole-4-carbaldehyde (4) [24] in ethanol
and in the presence of NaOH. Furthermore, condensation of
compound 1 with dimethylformamide dimethylaceal (6)
under reflux in dimethylformamide (DMF) according to the
reported method [25] afforded enaminone derivative 7
(Scheme 1).
Enaminones are stable compounds and constitute a
versatile class of useful precursors in organic synthesis, in
pharmaceutical development and in heterocyclic synthesis
[26]. Thus, treatment of enaminone derivative 7 with N-
nucleophile such as phenylhydrazine (8) in acetic acid under
reflux afforded pyrazole (9). The product was formed via
initial condensation of amino group of phenylhydrazine (7)
with carbonyl group of enaminone derivative 7 followed
by elimination of dimethylamine (10) (cf. Scheme 2) to give
the pyrazole 11, which is identical to that previously cited in
literature [26]. Similarly, transamination reaction between
enaminone (8) and 2-amino-2-(hydroxymethyl) propane-
1,3-diol (12) or p-phenylenediamine (13) in ethanol
produced 3-(1,3-dihydroxy-2-(hydroxylmethyl)propan-2-
ylamino)-1-(10H-phenothiazin-2-yl)prop-2-en-1-one (14)
and 3-(4-amino phenylamino)-1-(10H-phenothiazin-2-yl)
prop-2-en-1-one (15), respectively (Scheme 2).
Finally, condensation of acetyl derivative 1 with hydrazine
hydrate (19) or phenyhydrazine (8) gave the corresponding
hydrazones 35a and 35b. Vilsmeier–Haack reaction of
hydrazone derivative 35b with (POCl3/DMF) afforded the
target compound 3-(10H-phenothiazin-2-yl)-1-phenyl-1H-
pyrazole-4-carbaldehyde (36) (Scheme 6).
Biological evaluation
ABTS antioxidant assay.
The newly synthesized
compounds were screened for their antioxidant activity
using ABTS method [27].
Furthermore, the authors intended to introduce the 2H-
indazol-3(3aH)-one nucleus to the phenothiazine skeleton
in order to build a novel family of bioactive molecules.
Thus, Michael addition reaction of α,β-unsaturated ketone
5 with ethyl acetoacetate 16 in ethanol in the presence of so-
dium ethoxide gave the intermediate 17, which cyclized into
ethyl 2-oxo-4-(10H-phenothiazin-2-yl)-6-(1,3-diphenyl-1H-
pyrazol-4-yl) cyclohex-3-ene carboxylate (18) via Michael
addition followed by cyclocondensation. The behavior of
The results showed that most of the compounds
anchored to phenothiazine derivatives displayed good anti-
oxidant compared with ascorbic acid as shown in Table 1.
Bleomycin-dependent DNA damage. The bleomycin is a
family of glycopeptide antibiotics that was used routinely as
antitumor agents. The bleomycin assay was adopted for
assessing the pro-oxidant effects of food antioxidants. The
antitumor antibiotic bleomycin binds iron ions and DNA.
The bleomycin iron complex degrades DNA that upon
Scheme 1. Synthesis of α,β-unsaturated ketones 4, 5, and 6.
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Scheme 2. Synthesis of pyrazole 11 enaminones 14 and 15. [Color figure can be viewed in the online issue, which is available at wileyonlinelibrary.com.]
heating with thiobarbituric acid (TBA) yields a pink
chromogen. Upon the addition of suitable reducing agents,
antioxidants compete with DNA and diminish chromogen
formation [28], and the protective activity against DNA
damage induced by bleomycin iron complex was examined
for the new synthesized compounds. The results in Table 2
showed that compounds 11, 14, and 28 exhibited high
protection against DNA damage by comparing the results
obtained for the antioxidant properties of the compounds
reported in this study with their structures, and the following
structure–activity relationships were postulated in Figure 2
and indicate
(iv) Compounds 4 and 5 are lesss potent than compound
(1), which may be attributed to replacement of acetyl
group by α,β-unsaturated ketone.
EXPERIMENTAL
Instruments.
All melting points are recorded on
Gallenkamp electric melting point apparatus and are
uncorrected. IR spectra ν/cmÀ1 (KBr) were recorded on a
Mattson 5000 FTIR Spectrophotometer at Microanalytical
Center Faculty of Science, Mansoura University. 1H-
NMR and 13C-NMR spectra were measured on a Varian
Spectrophotometer at 300 and 75MHz, respectively,
using tetramethylsilane as an internal reference and
DMSO-d6 as a solvent. Mass spectra were recorded on
Kratos (70 EV) MS equipment and/or a Varian MAT
311A spectrometer at Microanalytical Center, Faculty of
Science, Cairo University. Elemental analyses (C and H)
were carried out at Microanalytical Center, Faculty of
Science, Cairo University; the results were found to be in
(i) Compound 20 has high antioxidant activity than 18,
which may be attributed to the presence of
bispyrazole moieties
(ii) Cyclization of the α,β-unsaturated ketones 4 or 5 into
pyrazole derivatives enhances the biological activity
of compounds 4 and 5.
(iii) Compound 28 has high antioxidant activities than
compound 6, which may be attributed to the presence
of pyrazole and thioamide moieties.
Scheme 3. Synthesis of pyrazoles 18, 20, and 22. [Color figure can be viewed in the online issue, which is available at wileyonlinelibrary.com.]
Journal of Heterocyclic Chemistry
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W. S. Hamama, M. A. Gouda, H. A. Kamal El-din, and H. H. Zoorob
Vol 000
Scheme 4. Synthesis of pyrazoles 23, 24, 25, and 26. [Color figure can be viewed in the online issue, which is available at wileyonlinelibrary.com.]
good agreement ( 0.3%) with the calculated values.
Reactions were monitored by thin-layer chromatography,
and preparative thin-layer chromatography carried out on
0.2- to 0.4-mm silica gel 60 F254 (Merck) plates using
UV light (254 and 366nm) for detection. Biological
testing was carried out at Drug Department, Faculty of
Pharmacy, Mansoura University, Egypt.
General procedure for the synthesis of (E)-1-(10H-
phenothiazin-8-yl)-3-phenylprop-2-en-1-one (4) and (E)-1-
(10H-phenothiazin-8-yl)-3-(1,3-diphenyl-1H-pyrazol-4-yl) prop-
2-en-1-one (5). 2-Acetylphenothiazine (1) (1g, 4mmol) in
absolute ethanol (50mL) was reacted with aldehydes,
namely, benzaldehyde (2) (0.43g, 4mmol) or 1,3-diphenyl-
1H-pyrazole-4-carbaldehyde (3) (0.99 g, 4mmol), then
NaOH (0.16mL, 10%) was added dropwise. The reaction
mixture was refluxed for 2 h. The resulting precipitate was
recrystallized from ethanol to give compounds 4 and 5,
respectively.
(4:1)]; IR (KBr): (ν/cmÀ1)= 3335 (NH), 1647 (CO),
1
1586cmÀ1 (C=C); H NMR (DMSO-d6) δ (ppm): 8.80 (s,
1H, NH), 7.96 (s, 1H, CHpyrazole), 7.69–6.76 (m, ArH,
17 H), 6.75 (d, J= 8.1 Hz, 1H, CO-CH=), 6.65 (d, J=8.1Hz,
1H, =CH); MS (EI, 70 ev) m/z (%)=473 (M++1, 21.0), 472
(M+, 100%), 252 (2.2), 251 (12.9), 245 (17.8), 241 (2.0),
226 (2.8), 219 (2.1), 198 (42.3), 77 (35.0); Anal. Calcd for
C30H21N3OS (471.57); Calcd: C: 76.41, H: 4.49; N, 8.91%.
Found: C: 76.33, H: 4.56; 8.85%.
Synthesis of (E)-3-(dimethylamino)-1-(10H-phenothiazin-8-
yl) prop-2-en-1-one (7). Compound 1 (1 g, 4 mmol) was
refluxed with N,N-dimethylformamide dimethyl acetal (6)
(0.5 mL, 4 mmol) in DMF (10 mL) for 10h. The formed
precipitate was filtered off and recrystallized from ethanol
to produce compound 7. Compound 1 was obtained as
orange precipitate in 92% yield; mp = 248–250°C;
Rf = 0.79 [pet. ether (40–60): ethyl acetate (4:2)]; IR
(KBr): (ν/cmÀ1)= 3256 (NH), 1636 (CO); 1H NMR
(DMSO-d6) δ (ppm): 2.93 (s, 3H, CH3); 3.11 (s, 3H,
CH3), 5.70 (d, 1H, =CH-CO, J = 12.4Hz), 6.64–7.35 (m,
7H, HAr), 7.72 (d, 1H, =CH-N, J= 12.3Hz), 8.70 (s, 1H,
NHphenothiazine); MS (EI, 70eV): m/z (%) =296.2 (M+,
100), 280.7 (4.7), 264.7 (8.5), 252.0 (22.2), 239.5 (1.1),
Compound 4 was obtained as red crystals in 72% yield;
mp= 199°C; [lit. ref. [24] mp: 199°C]; IR (KBr): (ν/cmÀ1
)
=3350(NH), 1650(CO), 1590 (C=C).
Compound 5 was obtained as red crystals in 69% yield;
mp= 222–221°C; Rf = 0.8 [pet. ether (40–60): ethyl acetate
Scheme 5. Synthesis of thiazoles 31 and 32 and oxazole derivative 34.
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Scheme 6. Synthesis of pyrazole carboxaldehyde 36. [Color figure can be viewed in the online issue, which is available at wileyonlinelibrary.com.]
226.7 (2.2), 197.4 (67.8), 99.2 (60.0), 71.0 (26.1), 55.1
(49.1); Anal. Calcd for C17H16N2OS (296.39); Calcd: C:
68.89; H: 5.44; N: 9.45%; Found: C: 68.83; H: 5.38; N:
Compound 15 was obtained as pale brown precipitate in
85% yield; mp >300°C; Rf = 0.58 [pet. ether (40–60): ethyl
acetate (4:2)]; IR (KBr): (ν/cmÀ1)=3471, 3361 (NH2, NH),
1
1
9.40%. Its IR, H NMR spectra, and MS spectrometry
1637 (CO), 1594 (C=C); H NMR (DMSO-d6) δ (ppm):
data are identical with that reported for compound 8 in
literature [29].
Synthesis of 2-(1-phenyl-1H-pyrazol-3-yl)-10H-phenothiazine
5.92 (d, 1H, CO-CH=, J= 7.8 Hz), 6.31 (d, 1H, =CH-CN,
J=12.0Hz), 6.53–7.86 (m, 11H, HAr), 8.71 (s, 1H,
NHphenothiazine), 12.03 (br.s, 3H, NH, NH2); MS (EI, 70eV):
m/z (%) =360 (M+ +1, 19.1), 359 (M+, 100), 344 (2.9), 279
(3.7), 198 (23.8), 161 (8.9), 145 (2.7), 120 (17.1), 107
(18.1), 92 (23.2); Anal. Calcd for C21H17N3OS (359.44): C:
70.17; H: 4.77; N: 11.69%; Found: C: 70.37; H: 4.47; N:
11.60%.
Synthesis of ethyl 2-oxo-4-(10H-phenothiazin-8-yl)-6-(1,3-
diphenyl-1H-pyrazol-4-yl)cyclohex-3-enecarboxylate (18). Ethyl
acetoacetate (16) (0.26g, 2mmol) was added to sodium
ethoxide solution that had been prepared from sodium
(0.044g, 2mmol) in absolute ethanol (50mL). After stirring
the reaction mixture for 1h, compound 5 (0.94g, 2mmol)
was added, and the reaction mixture was refluxed for 3h.
While still hot, the solution was added to a cold dilute
(11).
A solution of compound 7 (0.59g, 2mmol) and
phenylhydrazine (8) (0.2g, 2mmol) in glacial acetic acid
(50mL) was refluxed for 6h. The reaction mixture was
poured onto crushed ice. The formed solid was filtered off
and recrystallized from ethanol to produce 11 as gray
precipitate in 57% yield; mp= 106–108°C; Rf =0.35 [pet.
ether (40–60): ethyl acetate (4:2)]; IR (KBr): (ν/cmÀ1
)
1
=3378 (NH), 1597 (C=N), 1565 (C=C); H NMR (DMSO-
d6) δ (ppm): 6.53–7.74 (m, 14H, HAr), 8.64 (s, 1H,
NHphenothiazine); MS (EI, 70eV): m/z (%) = 343.1 (M+ +2,
100), 341.9 (M+, 6.8), 267.5 (3.4), 210.0 (6.8), 197.0 (3.4),
143.3 (3.4), 110.0 (3.4), 76.0 (3.4), 67.0 (8.5); Anal. Calcd
for C21H15N3S (341.43): C: 73.87; H: 4.43; N: 12.31%;
Found: C: 73.67; H: 4.37; N: 12.43%.
General procedure for the synthesis of (E)-3-(1,
3-dihydroxy-2-(hydroxymethyl) propan-2-ylamino)-1-(10H-
Table 1
phenothiazin-2-yl)prop-2-en-1-one
(14)
and
(E)-3-(4-
Antioxidant activity assay (ABTS) of some new
phenothiazine compounds.
aminophenylamino)-1-(10H-phenothiazin-2-yl)prop-2-en-1-
one (15). A solution of compound 7 (0.59 g, 2mmol) and
2-amino-2-(hydroxymethyl)propane-1,3-diol (12) (0.24g,
2 mmol) or benzene-1,4-diamine 13 (0.22 g, 2 mmol) in
absolute ethanol (50 mL) was refluxed for 6 and 3 h,
respoectively. The reaction mixture was poured on
crushed ice, and the separated solid was filtered off and
recrystallized from ethanol to give compounds 14 and 15,
respectively.
Inhibition
(%)
Compound no.
Absorbance of samples (λ)
Control of ABTSa
Ascorbic acid
0.516
0.038
0.152
0.190
0.366
0.229
0.098
0.092
0.112
0.121
0.089
0.100
0.137
0.102
0.176
0.091
0.100
0.105
0
88.54
70.25
63.17
29.06
55.62
81.29
81.81
78.62
76.08
83.01
80.91
73.85
80.53
65.21
82.01
80.23
79.24
1
5
6
7
Compound 14 was obtained as brown precipitate in 71%
yield; mp=250–252°C; Rf = 0.66 [pet. ether (40–60): ethyl ac-
etate (4:2)]; IR (KBr): (ν/cmÀ1)=3492 (OH), 3349 (NH),
10
12
16
19
26a
26b
36a
36b
40c
40d
43
48
1
1645 (CO), 1584 (C=C); H NMR (DMSO-d6) δ (ppm):
2.52 (s, 1H, NH), 2.81 (br.s 3H, 3OH), 3.30 (br.s, 6H,
3CH2), 5.63 (d, 1H, CO-CH=, J=12.0Hz), 6.60 (d, 1H,
=CH-CN, J=14.4Hz), 6.64–7.73 (m, 7H, HAr), 8.64 (s, 1H,
NHphenothiazine); MS (EI, 70 eV): m/z (%) =372 (M+, 0.5),
310 (4.9), 279 (100), 252 (42.6), 240 (17.6), 226 (11.4), 197
(86.4), 147 (10.6), 99 (84.8), 71 (36.4); Anal. Calcd for
C19H20N2O4S (372.44): C: 61.27; H: 5.41; N: 7.52%; Found:
C: 61.19; H: 5.48; N: 7.45%.
aABTS is the method used for antioxidant activity
(%) Inhibition = [A (control) À A (test)/A (control)] × 100.
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W. S. Hamama, M. A. Gouda, H. A. Kamal El-din, and H. H. Zoorob
Vol 000
Table 2
buff color crystals in 55% yield; mp=149–151°C;
crystallization solvent, ethanol; Rf = 0.62 [pet. ether (40–
60): ethyl acetate (4:1)]; IR (KBr): (ν/cmÀ1)=3449 (br,
Bleomycin-dependent DNA damage of the investigated compounds.
Compound no.
Absorbance of samples
1
2NH), 1735 (CO), 1639 (C=N), 1579 (C=C); H NMR
(DMSO-d6) δ (ppm):2.33 (m, 2H, CH2), 3.33 (m, 2H, CH),
6.60 (s, 1H, CH), 6.64–7.33 (m, 20H, HAr, NHphenothiazine),
8.60 (s, 1H, NHpyrazole); MS (EI, 70eV): m/z (%)=551
(M+, 13.1), 443 (20.6), 336 (0.3), 235 (35.4), 219 (29.4),
142 (0.4), 199 (2.9), 77.0 (81.1), 50 (100); Anal. Calcd for
C34H25N5OS (551.66): C: 74.02; H: 4.57; N: 12.70%;
Found: C: 73.91; H: 4.66; N: 12.83%.
Ascorbic acid
5
6
7
10
12
16
26a
26b
40d
43
48
0.062
0.112
0.137
0.128
0.088
0.058
0.095
0.064
0.073
0.079
0.088
0.094
Synthesis
of
(2Z)-ethyl-2-(hydroxyimino)-4-(10H-
phenothiazin-2-yl)-6-(1,3-diphenyl-1H-pyrazol-4-yl)cyclohex-3-
enecarboxylate (22). A mixture of compound 18 (0.58g,
1 mmol), hydroxylamine hydrochloride (0.07g, 1 mmol),
and sodium acetate (0.164 g, 2 mmol) in absolute ethanol
(25 mL) was refluxed for 6 h, then cooled and poured
onto ice to give solid precipitate, which recrystallized
from ethanol to furnish compound 22 as yellow solid
color in 50% yield; mp= 150–152°C; Rf = 0.63 [pet. ether
(40–60): ethyl acetate (4:1)]; IR (KBr): (ν/cmÀ1)= 3381
(OH), 3206 (NH), 1730 (COOEt), 1639 (C=N), 1598
hydrochloric acid. The red precipitate was isolated by
filtration, and then recrystallized from ethanol to afford 18.
Compound 18 was obtained as red crystals in 80% yield;
mp= 238–240°C; Rf = 0.61 [pet. ether (40–60): ethyl acetate
(4:1)]; IR (KBr): (ν/cmÀ1)= 3353 (NH), 1733 (COOEt),
1
1649 (br, CO, C=N), 1595 (C=C); H NMR (DMSO-d6) δ
(ppm): 1.13 (t, 3H, CH3), 2.49 (m, 2H, CH2), 3.07 (m, 1H,
CH), 3.33 (s, 1H, CH), 4.01 (q, 2H, COOCH2), 6.44 (s, 1H,
CH), 6.62–8.55 (m, 18H, HAr), 8.80 (s, 1H, NHphenothiazine);
13C NMR (DMSO-d6) δ (ppm): 193.6(CO), 169.2 (CO),
157.9, 151.0, 142.1 (2C), 139.4, 136.0, 132.8, 129.7 (2C),
128.5 (2C), 128.0 (2C), 127.7 (2C), 127.4 (2C), 126.3,
122.2, 122.0, 120.7, 120.1, 119.8, 117.9 (2C), 115.5, 114.5,
111.7, 60.1, 60.0, 35.9, 33.4, 13.5; MS (EI, 70eV): m/z (%)
=586 (M+ +3, 9.7), 555 (9.8), 538 (M+ -OEt,) 511.4 (6.5),
385 (5.6), 362 (9.7), 359 (92.5), 323 (19.9), 247 (100), 228
(1.9), 207 (1.8), 69 (9.9); Anal. Calcd for C36H29N3O3S
(583.70): C: 74.08; H: 5.01; N: 7.20%; Found: C: 74.13; H:
5.09; N: 7.12%.
1
(C=C); H NMR (DMSO-d6) δ (ppm): 1.14 (t, 3H, CH3),
2.43 (m, 2H, CH2), 3.24 (m, 2H, 2CH), 4.01 (q, 2H,
CH2), 6.63 (s, 1H, CH), 6.70–8.54 (m, 18H, HAr), 8.63
(s, 1H, NHphenothiazine), 11.21 (s, 1H, OH); MS (EI,
70eV): m/z (%) = 599 (M+ +1, 1.8), 598 (M+, 1.9), 580
(30.8), 551.3 (1.9), 511 (100), 508 (52.9), 346 (11.4),
212 (44.2), 198 (6.7), 105 (46.8), 77 (41.6); Anal. Calcd
for C36H30N4O3S (598.71): C: 72.22; H: 5.05; N: 9.36%;
Found: C: 71.92; H: 4.85; N: 9.21%.
General procedure for the synthesis of 1-(3-(10H-
phenothiazin-2-yl)-5-(1,3-diphenyl-1H-pyrazol-4-yl)-1H-pyrazol-
1-yl)ethanone (23) and 2-(1-phenyl-5-(1, 3-diphenyl-1H-pyrazol-
4-yl)-1H-pyrazol-3-yl)-10H-phenothiazine (24). A mixture of
compound (5) (0.47g, 1mmol) and hydrazine hydrate (19)
0.21g, 2mmol) or phenyl hydrazine (8) (0.21g, 2mmol) in
glacial acetic acid (5mL) was refluxed for 3h. The reaction
mixture was cooled and poured onto cold water (20mL). The
resulting solid was separated by filtration and recrystallized
from ethanol to give compounds 23 and 24, respectively.
Synthesis of 4,5-dihydro-6-(10H-phenothiazin-8-yl)-4-(1,3-
diphenyl-1H-pyrazol-4-yl)-2H-indazol-3(3aH)-one (20).
A
solution of compound 18 (0.58g, 1mmol) and hydrazine
hydrate (19) (0.1g, 1mmol) in glacial acetic acid (20mL)
was refluxed for 4h, then the reaction mixture was cooled
and poured onto crushed ice to give a precipitate, which
filtered off then recrystallized to furnish compound 20 as
Figure 2. Structure–activity relationship of the more potent antioxidant compounds.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2016
Bioactive Phenothiazine Derivatives
Compound 23 was obtained as pale yellow crystals in
79% yield; mp=134–136°C; Rf =0.56 [pet. ether (40–60):
ethyl acetate (4:1)]; IR (KBr): (ν/cmÀ1)=3390 (NH), 1644
acetate (4:1)]; IR (KBr): (ν/cmÀ1)=1684, 1654 (2CO,
C=N), 1596 (C=C); H NMR (DMSO-d6) δ (ppm): 2.13(s,
1
3H, CH3), 2.16 (s, 3H, CH3)), 7.36 (s, 1H, CHpyrazole 4 ‵),
7.38–8.03 (m, 17H, HAr), 8.26 (s, 1H, CHpyrazole 5 ‵‵). MS
(EI, 70eV): m/z (%)=569.8 (M+ +2, 17.1), 567.8 (M+, 4.1),
528 (100), 526 (33.5), 483 (63.9), 474 (17.9), 308 (20.4),
265 (20.2), 248 (11.8), 198 (29.0), 77 (66.2); Anal. Calcd
for C34H25N5O2S (567.66): C: 71.94; H: 4.44; N: 12.34%;
Found: C: 71.80; H: 4.34; N: 12.45%.
General procedure for the synthesis of 4, 5-dihydro-3-(10H-
phenothiazin-2-yl)-5-phenylpyrazole-1-carbothioamide (28) and
4, 5-dihydro-3-(10H-phenothiazin-2-yl)-5-(1, 3-diphenyl-1H-
pyrazol-4-yl)pyrazole-1-carbothioamide (29). A mixture of
arylidene 4 (0.33 g, 1 mmol) or (0.47g, 2mmol) 5 and
thiosemicarbazide 27 (0.1 g, 1.2 mmol) in absolute ethanol
(25mL) containing NaOH (2mL, 10%) was refluxed for
6 h. The reaction mixture was poured onto cold water
(30mL), and the formed precipitate was recrystallized from
ethanol to give the corresponding pyrazole derivatives 28
and 29.
1
(CO, C=N), 1598 (C=C); H NMR (DMSO-d6) δ (ppm):
2.24 (s, 3H, COCH3), 6.60 (s, 1H, CHpyrazole4 ‵), 6.65–7.84
(m, 17H, HAr), 8.33 (s, 1H, CHpyrazole5 ‵‵), 8.72 (s, 1H,
NHphenothiazine); MS (EI, 70eV): m/z (%)=528 (M+ +2,
91.7), 525 (M+, 10.3), 486 (20.5), 473 (100), 307 (4.2),
265 (12.4), 28 (70.5), 109 (17.4); Anal. Calcd for
C32H23N5OS (525.62): C: 73.12; H: 4.41; N: 13.32%;
Found: C, 73.20; H, 4.32; N: 13.22%.
Compound 24 was obtained as yellow crystals in 84%
yield; mp=110–112°C; Rf = 0.86 [pet. ether (40–60): ethyl
acetate (4:1.5)]; IR (KBr): (ν/cmÀ1)=3386 (NH), 1620
1
(C=N), 1569 (C=C); H NMR (DMSO-d6) δ (ppm): 6.65
(s, 1H, CHpyrazole 4 ‵), 6.76–7.94 (m, 22H, HAr), 8.33 (s,
1H, CHpyrazole 5 ‵‵), 8.83 (s, 1H, NHphenothiazine); 13C NMR
(DMSO-d6) δ (ppm): 150.3, 147.1, 143.4, 142.1, 139.3
(2C), 135.7, 132.7 (2C), 132, 131.5, 129.7 (2C), 129.6
(2C) 129.5 (2C), 128.3 (2C), 127.9 (2C), 127.6, 126.5
(2C), 121.8 (2C),121.7(2C) 119.1, 118.9, 118.3 (2C),
116.3 (2C), 106.7 (2C); MS (EI, 70eV): m/z (%)=561.0
(M+ +1, 14.9), 559.9 (M+, 64.8), 528 (100), 483 (45.4),
341 (38.9), 264 (12.4), 224 (18.6), 219 (9.5), 197 (12.1),
77 (58.1); Anal. Calcd for C36H25N5S (559.68): C: 77.26;
H: 4.50; N: 12.51%; Found: C: 77.17; H: 4.76; N: 12.59%.
2-(5-(1,3-Diphenyl-1H-pyrazol-4-yl)-1H-pyrazol-3-yl)-10H-
Compound 28 was obtained as yellow crystals in 81%
yield; mp = 220–221°C; Rf = 0.33 [pet. ether (40–60): ethyl
acetate (4:1)]; IR (KBr): (ν/cmÀ1)= 3414, 3350, 3264
1
(NH2, NH), 1659 (C=N), 1582 (C=C); H NMR (DMSO-
d6) δ (ppm): 3.13 (s, 2H, CHpyrazole 4‵), 4.08 (t, 1H,
CHpyrazole 5 ‵), 5.66 (s, 2H, NH2), 6.67–7.70 (m, HAr,
12H), 8.82 (s, 1H, NHphenothiazine); MS (EI, 70eV): m/z
(%) = 402.5 (M+, 52.4), 326 (13.4), 265 (19.4), 224 (100),
204 (14.4), 198 (84.5), 145 (4.4), 127 (14.4), 77 (25.7),
61 (9.4); Anal. Calcd for C22H18N4S2 (402.54): C: 65.64;
H: 4.51; N: 13.92%; Found: C: 65.49; H: 4.43; N: 13.95%.
Compound 29 was obtained as dark yellow crystals in
75% yield; mp= 169–170°C; Rf = 0.28 [pet. ether (40–
60): ethyl acetate (4:1)]; IR (KBr): (ν/cmÀ1) = 3424,
3334, 3250 (NH2, NH), 1623 (C=N), 1597 (C=C); 1H
NMR (DMSO-d6) δ (ppm): 3.08 (s, 2H, CHpyrazole 4‵),
4.09 (t, 1H, CHp, yrazole5), 6.54 (s, 2H, NH2), 6.63–7.91
(m, 18H, HAr), 8.61 (s, 1H, NHphenothiazine); MS (EI,
70eV): m/z (%) =544.4 (M+, 0.6), 324 (3.1), 285 (13.6),
269 (12.9), 263 (33.4), 198 (20.1), 136 (57.7), 124
(55.9), 97 (100), 58 (17.4); Anal. Calcd for C31H24N6S2
(C, 68.36; H, 4.44; N, 15.43; Found: C: 68.30; H: 4.35;
N: 15.34%.
General procedure the for synthesis of 2-(4,5-dihydro-5-
phenyl-1-(4-phenylthiazol-2-yl)-1H-pyrazol-3-yl)-10H-
phenothiazine (31) and 2-(4,5-dihydro-5-(1,3-diphenyl-1H-
pyrazol-4-yl)-1-(4-phenylthiazol-2-yl)-1H-pyrazol-3-yl)-10H-
phenothiazine (32). A mixture of compound 28 (0.40 g,
1 mmol) or compound 29 (0.54g, 1 mmol) with phenacyl
bromide (30) (0.2 g, 1 mmol) in absolute ethanol (25 mL)
was refluxed for 6 h, then allowed to cool and poured onto
cold water (20 mL). The resulting solid was collected and
recrystallized from ethanol to give the corresponding
pyrazolylthiazoles 31 and 32.
phenothiazine (25).
A solution of compound 5 (0.94g,
2mmol) and hydrazine hydrate (19) (0.3g, 3mmol) in
absolute ethanol (10mL) was refluxed for 3h. The
reaction mixture was diluted with cold water (20mL). The
formed solid was separated by filtration and recrystallized
to give 25.
Compound 5 was obtained as buff crystals in 87% yield;
mp=150–151°C; crystallization solvent, ethanol; Rf =0.53
[pet. ether (40–60): ethyl acetate (4:1)]; IR (KBr): (ν/cmÀ1
)
= 3387 (NH), 1623 (C=N), 1551 (C=C); 1H NMR
(DMSO-d6) δ (ppm): 6.61 (s, 1H, NHpyrazole 1 ‵), 6.66 (s,
1H, CHpyrazole 4 ‵), 6.73–7.92 (m, 17H, HAr), 8.43 (s, 1H,
CHpyrazole 5 ‵‵), 8.53 (s, 1H, NHphenothiazine); MS (EI, 70eV):
m/z (%)=484.6 (M+ +1, 3.2), 483.5 (M+, 10.2), 457.2
(100), 264 (1.6), 236 (35.3), 224 (15.7), 219 (4.7), 198
(16.5), 77 (31.3), 65 (1.8); Anal. Calcd for C30H21N5S
(483.59): C: 74.51; H: 4.38; N: 14.48%; Found: C: 74.40;
H: 4.30; N: 14.36%.
1-(3-(N-Acetyl-phenothiazin-2-yl)-5-(1,3-diphenyl-1H-pyrazol-4-
yl)-1H-pyrazol-1-yl)ethanone (26). A solution of compound 23
(0.48g, 1mmol) was refluxed for 3h in a mixture of glacial
acetic acid/acetic anhydride (6mL, 1:1 v). The reaction
mixture was diluted with cold water (20mL). The resulting
solid was separated by filtration and recrystallized from
ethanol to give the desired pyrazole derivative 26.
Compound 23 was obtained as yellow crystals in 70%
yield; mp= 216–217°C; Rf =0.35 [pet. ether (40–60): ethyl
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

W. S. Hamama, M. A. Gouda, H. A. Kamal El-din, and H. H. Zoorob
Vol 000
Compound 31 was obtained as green crystals in 85%
yield; mp= 260–262°C; Rf =0.83 [pet. ether (40–60): ethyl
acetate (4:1)]; IR (KBr): (ν/cmÀ1) =3441 (NH), 1630
66.08; H: 4.80; N: 16.33; MS (EI, 70eV): m/z (%)
= 255.3 (M+, 100), 240 (22.2), 224 (35.7), 198 (45.5),
166 (8.6), 146 (2.0), 121 (11.1), 104 (9.3), 77 (11.4), 58
(3.9%.); Anal. Calcd for C14H13N3S (255.34): C: 65.85;
H: 5.13; N: 16.46%.
Compound 35b was obtained as yellow crystals in 80%
yield; mp=253À254°C; Rf = 0.62 [pet. ether (40–60): ethyl
acetate (4:1)]. Its IR, 1H NMR, and MS spectral data are iden-
tical with that reported for compound 35b in literature [30].
3-(10H-Phenothiazin-2-yl)-1-phenyl-1H-pyrazole-4-
1
(C=N), 1545 (C=C); H NMR (DMSO-d6) δ (ppm):3.11
(s, 2H, CHpyrazole 4‵), 4.05 (t, 1H, CHpyrazole 5 ‵), 6.93–
7.95 (m, 19H, HAr), 8.63 (s, 1H, NHphenothiazine); MS (EI,
70eV): m/z (%)= 502.2 (M+, 86.4), 425 (4.3), 341 (11.4),
304 (2.6), 224 (100), 198 (43.3), 160 (4.6), 153 (23.8);
Anal. Calcd for C30H22N4S2 (502.65): C: 71.68; H: 4.41;
N: 11.15%; Found: C: 71.53; H: 4.57; N: 11.24%.
Compound 32 was obtained as dark green crystals in 79%
yield; mp=240–241°C; Rf =0.34 [pet. ether (40–60): ethyl
acetate (4:1)]; IR (KBr): (ν/cmÀ1)=3433 (NH), 1623
carbaldehyde (36).
To a stirred solution of compound
35b (1 g, 3mmol) in DMF (5mL) was added dropwise
phosphorus oxychloride (POCl3) (1 mL) through (10–
15min), then left while stirring for 2h at room
temperature, followed by reflux on steam bath for 2 h and
poured onto crushed ice. Stirring was continued while
adding sodium bicarbonate until the effervescence ceased
to give a precipitate of 36. Compound 36 was obtained as
buff crystals in 62% yield; mp = 150–151°C;
crystallization solvent, ethanol; Rf = 0.43 [pet. ether (40–
60): ethyl acetate (4:1)]; IR (KBr): (ν/cmÀ1) =3343 (NH),
1
(C=N), 1530 (C=C); H NMR (DMSO-d6) δ (ppm):3.13
(s, 2H, CHpyrazole 4‵), 4.07 (t, 1H, CHpyrazole 5 ‵), 6.95–7.83
(m, 24H, HAr), 7.95 (s, 1H, NHphenothiazine); MS (EI,
70eV): m/z (%)=644.4 (M+, 2.0), 224 (28.3), 219 (4.5),
217 (16.9), 198 (17.3), 160 (7.1), 150 (18.5), 134.3 (30.2),
105 (100), 83 (32.1); Anal. Calcd for C39H28N6S2
(644.81): C: 72.64; H: 4.38; N: 13.03%; Found: C: 72.53;
H: 4.45; N: 12.90%.
1
2-(5-(1,3-Diphenyl-1H-pyrazol-4-yl)isoxazol-3-yl)-10H-
1666 (CO), 1610 (C=N), 1563 (C=C); H NMR (DMSO-
phenothiazine (34).
A solution of hydroxylamine
d6) δ (ppm): 6.64–8.08 (m, 13H, HAr), 8.75 (s, 1H, NH
phenothiazine), 11.22 (s, 1H, CHO); MS (EI, 70eV): m/z
(%) = 369 (M+, 7.7), 341 (100), 297 (4.4), 274 (13.1),
264 (9.1), 198 (70.6), 171 (20.2), 93 (9.4), 77 (32.4), 65
(7.3); Anal. Calcd for C22H15N3OS (369.44): C: 71.52;
H: 4.09; N: 11.37%; Found: C: 71.35; H: 4.16; N: 11.31%.
Pharmacological studies. More details about the ABTS
method antioxidant assay [27] and Bleomycin-dependent
DNA damage method [31,32] were presented in literature
[33].
hydrochloride 33 (0.069 g, 1 mmol) and sodium acetate
(0.16 g, 2 mmol) in a least amount of water was added to
a suspension of 5 or (0.47 g, 1 mmol) in absolute ethanol
(50 mL). The reaction mixture was refluxed for 4 h, then
cooled and poured onto water (15 mL). The obtained solid
was recrystallized from ethanol to give 34 as dark yellow
in 89% yield; mp = 120–122°C; Rf = 0.40 [pet. ether (40–
60): ethyl acetate (4:1.5)]; IR (KBr): (ν/cmÀ1) = 3382
(NH), 1620 (C=N), 1598 (C=C), 1501 (NO); MS (EI,
70eV): m/z (%) = 487.0 (M+ +3, 10.8), 486 (M+ +2, 6.9),
484 (M+, 15.4), 264 (14.5), 256 (100), 244 (29.3), 224
(73.9), 219 (21.6), 198 (98.4), 69 (11.0); 1H NMR
(DMSO-d6) δ (ppm): 7.92–6.65 (m, 19H, HAr), 8.64 (s,
1H, NHphenothiazine); Anal. Calcd for C30H20N4OS
(484.57): C: 74.36; H: 4.16; N: 11.56%. Found: C: 74.22;
H: 4.25; N: 11.45%.
General procedure for the synthesis of 2-(1-
hydrazonoethyl)-10H-phenothiazine (35a) and 1-(1-(10H-
Phenothiazin-2-yl)ethylidene)-2-phenylhydrazine (35b). 2-
Acetylphenothiazine (1) (0.5 g, 2mmol) and hydrazine
hydrate (19) (0.22 g, 2 mmol) or phenylhydrazine (8)
(0.22 g, 2 mmol) in ethanol (20 mL) was refluxed for 6h
and 20min, respectively. The formed precipitate was
filtered off and crystallized from ethanol to give
compounds 35a and 35b, respectively.
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Journal of Heterocyclic Chemistry
DOI 10.1002/jhet