An efficient one-pot reaction for the synthesis of pyrazolones bearing a phenothiazine unit

Article abstract of DOI:10.1016/j.tetlet.2012.08.152

A simple one-pot approach for the synthesis of new pyrazolones containing a phenothiazine unit is reported. Compound 4b was evaluated for its antiproliferative activity on a NCI-60 cancer cell line panel and exhibited selective cytostatic activity toward CCRF-CEM, HL-60(TB), K-562, MOLT-4, and RPMI-8226 leukemia cell lines.

Full text of DOI:10.1016/j.tetlet.2012.08.152

Tetrahedron Letters 53 (2012) 6127–6131
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Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
An efficient one-pot reaction for the synthesis of pyrazolones bearing
a phenothiazine unit
Lavinia Baciu-Atudosie a, Alina Ghinet a,b,c, Dalila Belei a, Philippe Gautret b,c, Benoît Rigo b,c, Elena Bîcu a,
⇑
a Department of Organic Chemistry, ‘Al. I. Cuza’ University of Iasi, Faculty of Chemistry, Bd. Carol I nr. 11, 700506 Iasi, Romania
b Univ Lille Nord de France, F-59000 Lille, France
c UCLille, EA 4481 (GRIIOT), Laboratoire de Pharmacochimie, HEI, 13 rue de Toul, F-59046 Lille, France
a r t i c l e i n f o
a b s t r a c t
Article history:
A simple one-pot approach for the synthesis of new pyrazolones containing a phenothiazine unit is
reported. Compound 4b was evaluated for its antiproliferative activity on a NCI-60 cancer cell line panel
and exhibited selective cytostatic activity toward CCRF–CEM, HL-60(TB), K-562, MOLT-4, and RPMI-8226
leukemia cell lines.
Received 20 July 2012
Revised 20 August 2012
Accepted 31 August 2012
Available online 7 September 2012
Ó 2012 Elsevier Ltd. All rights reserved.
Keywords:
Pyrazolone
Phenothiazine
One-pot reaction
Hydrazine
Heterocyclic compounds with a pyrazolone skeleton occupy a
significant place both in medicinal chemistry and in organic syn-
thesis. In the field of medicinal chemistry, they have been investi-
gated for use as cardiotonics,1 for their potential antitumor
activity,2–4 as potent inhibitors of the enzyme GSK3b,5 of tubulin
procedures21–23 (Scheme 1 and Table 1), free N-substituted hydra-
zine 6 was not obtained. On heating a solution of 10-(chloroace-
tyl)-10H-phenothiazine (5) and hydrazine hydrate (molar ratio of
1:1.521 or 1:4,22 respectively) in absolute ethanol, we isolated, as
unique reaction products, hydrazones 724 and 8,25 and the cleavage
product, phenothiazine 9.
polymerization,6 or of TNF-
a
production7 (Fig. 1). The pyrazolone
moiety is also found in the structure of antibacterial and antifungal
agents,8 anticonvulsant compounds,9 and molecules with antiviral
activity against PPRV (peste des petits ruminant virus).10 These
structures also demonstrate insecticidal activity against Tetrany-
chus cinnabarinus,11 have furnished novel antituberculars,12 and
represent promising starting materials for lead optimization aimed
at the discovery of DMT1 blockers as iron overload therapeutics13
and antioxidants.14 In the field of organic synthesis, pyrazolones
are important intermediates for the synthesis of dihydro[2,3-c]pyr-
azoles,15–18 phenylpyrazoles,19 and chromeno[2,3-c]pyrazol-
4(1H)-ones.20
Employing alternative literature conditions (benzene, reflux),23
we isolated the hydrochloride salt 10, which was the expected
product. Formation of the same salt starting from hydrazine 6,
which was obtained with an optimized reaction described in Ta-
ble 2 (entries 3 and 4), and hydrochloric acid in methanol, fur-
nished a chemical proof of this structure.
Due to the instability of hydrazine 6 when heated in ethanol
(see below), we undertook the optimization of its synthesis at a
lower temperature (5 °C) using chloroacetyl compound 5 in DMA,
absolute EtOH, and [bmim][BF4]/CH3CN respectively, in the pres-
ence of excess of hydrazine hydrate (Table 2). The best yields were
obtained when using DMA in the presence of 4 Å molecular sieves
(entries 3 and 4).26
The objective of our work consisted of the synthesis of new pyr-
azolone derivatives 4 with potential antitumor properties, substi-
tuted by
a
2-oxoethylphenothiazine moiety (Fig. 1). These
It was observed in the initial reactions (Scheme 1) that hydra-
zine 6 was rapidly oxidized to hydrazone 7. Chemical evidence
for the formation of compound 7 was provided by its reaction with
ethyl acetoacetate (11) (Scheme 2) which, conducted in DMA at
80 °C, led to the formation of enamino ester 13 in 63% yield.27
While investigating the mechanism of the formation of hydra-
zone 7, it was observed that a Wolff–Kishner reaction occurred
(without base) when the hydrazine 6 was refluxed in ethanol for
24 h, and known28 10-acetyl-10H-phenothiazine (15) was isolated
compounds were obtained via a one-pot reaction of reactive halo-
genated reagents, hydrazine hydrate, and b-ketoesters.
Initially, a stepwise approach was investigated for the synthesis
of pyrazolones 4a–d starting with the reaction of N-chloroacetyl
compound 5 with hydrazine hydrate. However, using literature
⇑
Corresponding author.
E-mail address: elena@uaic.ro (E. Bîcu).
0040-4039/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetlet.2012.08.152

6128
L. Baciu-Atudosie et al. / Tetrahedron Letters 53 (2012) 6127–6131
Table 1
H
H
O
N
N
Results obtained by our group versus the results described in the literature
OH
O
Ratio
5:H4N2ÁH2O
1:1.5
Reaction conditions
Producta Mp (°C)
Lit. mpb
(°C)
O
O
214–215 180c
O
Absolute EtOH, reflux, 18 h
7
8
7
230–232
N
214–215 145d
N
1:4
1:1
Absolute EtOH, reflux, 5 h
Benzene, reflux, 3 h
O
e,f
N
10
148–152
—
N
H
H
O
a
b
c
d
e
f
O
Isolated products.
Melting points related in the literature for the final products.
1
2
Lit. 6
Lit. 5
Lit.21
Lit.22
2
Not described.
R
N
Lit.23
N
O
N
N
O
N
HN
O
N
Table 2
1
Optimization of the reaction conditions for the synthesis of 10-(hydrazinoacetyl)-
10H-phenothiazine (6) at 5 °C
R
N
S
Entry Solvent
Ratio 5:
H4N2ÁH2O
Time
(h)
Yield
(%)
4
3 Lit. 7
1
2
3
DMA
1:2
1:4
1:2
5
4
5
37
50
56
Figure 1. Structures of known pyrazolones 1, 2, and 3 with antitumor potential, and
of the target pyrazolones 4 with a phenothiazine unit.
DMA, 4 Å molecular
sieves
4
5
6
7
8
1:4
1:2
1:4
1:2
1:4
4
3
3
5
5
58
18
22
42
48
in 90% yield29 (Scheme 3). However, when the reaction was con-
ducted at room temperature for 24 h, hydrazone 7 was obtained
quantitatively. It should also be noted that very few references re-
port the instability of N-carbonylmethylhydrazines in the presence
of air or describe the necessity to work in deoxygenated condi-
tions.30a–c We previously observed oxidations promoted by oxygen
from air dissolved in alcohols.30d–g
Absolute EtOH
[bmim][BF4]/MeCN
and 4b (Scheme 4).31 It was reported that some pyrazoles can only
be obtained in the presence of catalysts.32 In the case of hydrazine
6 and ketoester 11 (DMA, 110 °C), the main product was the
Having optimized the method for the generation of hydrazine 6,
we next studied the synthesis of the corresponding pyrazolones 4a
NH2
HN
O
S
N
Lit. 21-23
N
O
i, ii or iii
S
6
Cl
NH2
NH
O
N
N
O
O
N
H
N
N
N
S
i
+
+
S
ii
5
S
S
7
21%
54%
8
5%
0%
9
69%
46%
.
Our results
NH2 HCl
HN
O
N
S
iii
10 72%
Scheme 1. Reagents and conditions described in the literature for the synthesis of hydrazine 6, and our results under the same conditions: (i) 10-(chloroacetyl)-10H-
phenothiazine (5) (1 equiv), hydrazine hydrate (1.5 equiv), absolute EtOH, reflux, 18 h, (compound 6, yield not specified21); (ii) 5 (1 equiv), hydrazine hydrate (4 equiv),
absolute EtOH, reflux, 5 h (compound 6, 85%22); (iii) 5 (1 equiv), hydrazine hydrate (1 equiv), benzene, reflux, 3 h (compound 6, yield not specified23).

L. Baciu-Atudosie et al. / Tetrahedron Letters 53 (2012) 6127–6131
6129
COOEt
COOEt
NH
N
N
NH2
N
N
OEt
O
O
O
O
O
N
S
N
N
11
i
S
S
13 63%
12
7
Scheme 2. Synthesis of compound 13. Reagents and conditions: (i) hydrazone 7 (1 equiv), ethyl acetoacetate (11) (2 equiv), DMA, 80 °C, 24 h.
NH2
NH
NH2
N
N
HN
O
O
O
O
N
N
N
N
ii
i
- N2
- H2O
S
S
S
S
7
6
14
15
Scheme 3. Decomposition of 10-(hydrazinoacetyl)-10H-phenothiazine (6). Reagents and conditions: (i) EtOH, rt, 24 h, quantitative; (ii) EtOH, reflux, 24 h, 90%.
R2
N
R2
N
NH2
O
OEt
N
N
HN
O
O
O
N
S
N
S
N
O
O
i
+
+
2
R
OEt
S
4a R2 = Me 14%
4b R2 = Ph 73%
17 R2 = Me 62%
18 R2 = Ph 0%
6
11 R2 = Me
16 R2 = Ph
Scheme 4. Standard method for the synthesis of pyrazolones 4a33 and 4b.36 Reagents and conditions: (i) hydrazine 6 (1 equiv), b-ketoester 11 or 16 (1 equiv), DMA, 110 °C,
24 h.
ethoxypyrazole 17.34 Formation of this compound was unex-
pected; however, analogous reactions have already been
reported.32,35
Taking all this information into account, the pyrazolone ring
closure via a one-pot reaction was realized under mild conditions
(Scheme 5). Thus, a solution of hydrazine hydrate in DMA was
cooled to 0–5 °C, and then the halogenated derivatives 5, 19, or
20 was added and the mixture was stirred at this temperature until
complete consumption of the chloroacetyl derivative. Upon addi-
tion of b-ketoester 11 or 16, the mixture was reacted for 3 h at
2
R
N
O
N
Cl
O
O
1
1
N
R
N
S
R
O
O
i
.
+
+ H2N NH2 H2O
2
R
OEt
S
4a R1 = H, R2 = Me 65%
4b R1 = H, R2 = Ph 62%
4c R1 = Cl, R2 = Me 59%
4d R1 = CF3, R2 = Me 58%
11 R2 = Me
16 R2 = Ph
5
R1 = H
19 R1 = Cl
20 R1 = CF3
Scheme 5. One-pot method for the synthesis of pyrazolones 4a–d.37 Reagents and conditions: (i) hydrazine (2 equiv), b-ketoester 11 or 16 (1 equiv), 4 Å molecular sieves
powdered, DMA, 0 ? 60 °C, 24 h.

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L. Baciu-Atudosie et al. / Tetrahedron Letters 53 (2012) 6127–6131
Figure 2. Results of the in vitro human cell growth inhibition obtained with pyrazolone 4b at a 10 l
M concentration.38
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room temperature, and then heated for 8 h at 60 °C. Pyrazolones
4a–d were thus obtained in good yields (Scheme 5), without any
traces of the corresponding ethoxypyrazoles being evident.
Pyrazolone 4b was selected by the NCI (National Cancer Insti-
tute, USA) for screening against 60-human tumor cell lines and
showed selective growth inhibition toward leukemia cells at a
10 lM concentration (73% inhibition of CCRF–CEM, 23% inhibition
of HL-60(TB), 26% inhibition of K-562, 45% inhibition of MOLT-4,
and 16% inhibition of RPMI-8226) (Fig. 2). These results indicate
that pyrazolone derivatives bearing a phenothiazine unit are
potentially good starting points for further investigation as new
anticancer agents.
In conclusion, we have developed an efficient and simple one-
pot reaction for the synthesis of pyrazolones bearing a phenothia-
zine unit. This synthetic strategy leads easily to the target
compounds in higher yields than the standard two-step method.
The synthesis of 10-(hydrazinoacetyl)-10H-phenothiazine (6) was
realized and optimized. This optimization study permitted the
isolation and characterization of new compounds with a phenothi-
azine core.
22. Salvi, V. K.; Sharma, S.; Sharma, C.; Bhambi, D.; Talesara, G. L. Indian J. Chem.
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23. Rajasekaran, A.; Periasamy, M.; Venkatesan, S. J. Dev. Biol. Tissue Eng. 2010, 2, 5–
13.
Acknowledgements
24. (1E)-Oxo-(10H-phenothiazin-10-yl)acetaldehyde hydrazone (7). Compound
obtained using the same reaction conditions as described in the
literature,21,22 after purification by column chromatography on silica gel
using EtOAc/n-hexane 8/2 as eluent. White solid, 21% yield; mp 214–215 °C. IR
The authors gratefully acknowledge the ‘Programul Operational
Sectorial Dezvoltarea Resurselor Umane 2007–2013’ (project POS-
DRU/88/1.5/S/47646) and the CommScie (project POSDRU/89/1.5/
S/63663) for financial support, and the NCI (National Cancer Insti-
tute) for the biological evaluation of compound 4b on their 60-cell
panel.
(ATR) (m
max/cmÀ1): 3436, 3311, 3155, 3062, 1631, 1517, 1459, 1420, 1316,
1233, 1129, 757. 1H NMR (DMSO-d6, 400 MHz) (d ppm) 7.20 (s, 1H), 7.31 (t,
J = 7.6 Hz, 2H), 7.41 (t, J = 7.6 Hz, 2H), 7.56 (d, J = 7.6 Hz, 2H), 7.61 (d, J = 7.6 Hz,
2H), 8.21 (s, 2H). 13C NMR (DMSO-d6, 100 MHz) (d ppm) 124.8 (CH), 126.8
(2CH), 127.6 (2CH), 127.3 (2CH), 127.8 (2CH), 131.7 (2C), 138.4 (2C), 161.3 (C).
MS ESI m/z (rel%): 269.1 (M+, 7), 199 (100), 198 (72), 167 (25), 241 (15).
25. (1E)-Oxo(10H-phenothiazin-10-yl)acetaldehyde[2-oxo-2-(10H-phenothiazin-10-
yl)ethyl]hydrazone (8). By-product obtained using the same reaction conditions
as described in the literature,21 after purification by column chromatography
on silica gel using EtOAc/n-hexane 8/2 as eluent. White solid, 5% yield; mp
References and notes
230–232 °C. IR (ATR) (m
max/cmÀ1): 1674, 1524, 1461, 1279, 1257, 752. 1H NMR
1. Jarreau, F.X.; Koenig, J.J. U.S. Patent 4526,895, 1985. Chem. Abstr. 1996, 97,
225,868/20.
(CDCl3, 400 MHz) (d ppm) 4.32 (br s, 2H), 6.69 (s, 1H), 7.20 (t, J = 7.6 Hz, 2H),
7.24 (t, J = 7.6 Hz, 2H), 7.26–7.35 (m, 4H), 7.41 (d, J = 7.6 Hz, 2H), 7.44 (d,
J = 7.6 Hz, 2H), 7.53 (d, J = 7.6 Hz, 2H), 7.59 (d, J = 7.6 Hz, 2H), 11.29 (t,
J = 4.4 Hz, 1H). 13C NMR (CDCl3, 100 MHz) (d ppm) 53.5 (CH2), 117.9 (CH),
126.8 (3CH), 126.9 (3CH), 127.1 (2CH), 127.1 (2CH), 127.2 (2CH), 127.9 (2CH),
128.1 (2CH), 132.7 (2C), 133.1 (2C), 137.7 (2C), 137.8 (2C), 161.3 (C), 168.5 (C).
MS ESI m/z (rel%): 508 (M+, 3), 198 (100), 254 (19), 167 (17).
2. Farghaly, A. R. Arkivoc 2010, xi, 177–187.
3. Zhang, Y.; Zhang, L.; Liu, L.; Guo, J.; Wu, D.; Xu, G.; Wang, X.; Jia, D. Inorg. Chim.
Acta 2010, 363, 289–293.
-
4. Markovic´, V.; Eric, S.; Stanojkovic´, T.; Gligorijevic´, N.; Arandelovic´, S.;
´
´
´
´
´
Todorovic, N.; Trifunovic, S.; Manojlovic, N.; Jelic, R.; Joksovic, M. D. Bioorg.
Med. Chem. Lett. 2011, 21, 4416–4421.
5. Arnost, M.; Pierce, A.; Haar, E.; Lauffer, D.; Madden, J.; Tanner, K.; Green, J.
Bioorg. Med. Chem. Lett. 2010, 20, 1661–1664.
26. 10-(Hydrazinoacetyl)-10H-phenothiazine (6). Chloroacetylphenothiazine
5
(10 mmol) in DMA (12 mL) containing molecular sieves 4 Å (0.8 g) was
cooled to 0 °C. Hydrazine hydrate was then added (40 mmol) and the
resulting reaction media were stirred at 5 °C for 4 h. A mixture ice/distilled
H2O was added to the final suspension. The resulting solid was collected by
filtration and purified by column chromatography on silica gel using CHCl3/
ˇ
ˇ
6. Burja, B.; Cimbora-Zovko, T.; Tomic´, S.; Jelušic´, T.; Kocevar, M.; Polanc, S.;
Osmak, M. Bioorg. Med. Chem. 2010, 18, 2375–2387.
7. Braña, M. F.; Gradillas, A.; Ovalles, A. G.; López, B.; Acero, N.; Llinares, F.; Muñoz
Mingarro, D. Bioorg. Med. Chem. 2006, 14, 9–16.

L. Baciu-Atudosie et al. / Tetrahedron Letters 53 (2012) 6127–6131
6131
Ketone form (4a): 1H NMR (CDCl3, 400 MHz) (d ppm) 2.06 (s, 3H), 3.21 (s, 2H),
4.57 (br s, 2H), 7.25 (t, J = 7.6 Hz, 2H), 7.33 (t, J = 7.6 Hz, 2H), 7.45 (d,
J = 7.6 Hz, 2H), 7.57 (d, J = 7.6 Hz, 2H). 13C NMR (CDCl3, 100 MHz) (d ppm)
17.0 (CH3), 41.0 (CH2), 46.6 (CH2), 126.8 (2CH), 127.2 (2CH), 127.3 (2CH),
128.1 (2CH), 133.0 (2C), 137.6 (2C), 155.8 (C), 165.7 (C), 173.4 (C). MS ESI m/z
(rel%): 337 (M+, 7), 199 (100), 198 (38), 167 (34), 111 (32). Phenolic form
(4a’): 1H NMR (DMSO-d6, 400 MHz) (d ppm) 1.98 (s, 3H), 4.71 (br s, 2H), 5.09
(s, 1H), 7.10–8.00 (m, 8H), 10.85 (br s, 1H). 13C NMR (DMSO-d6, 100 MHz) (d
ppm) 14.0 (CH3), 48.0 (CH2), 85.0 (CH), 127.2 (2CH), 127.4 (4CH), 128.0 (2CH),
132.0 (2C), 137.5 (2C), 146.0 (C), 153.8 (C), 165.7 (C).
EtOH 8/2 to afford pure hydrazine 6 as a white solid, 72% yield; mp 95–98 °C. IR
(ATR) (
max/cmÀ1): 3000–3350, 1679, 1459, 1381, 1252, 767. 1H NMR (DMSO-
m
d6, 400 MHz) (d ppm) 3.30–3.70 (br s, 5H), 7.33 (t, J = 7.6 Hz, 2H), 7.41 (t,
J = 7.6 Hz, 2H), 7.57 (d, J = 7.6 Hz, 2H), 7.64 (d, J = 7.6 Hz, 2H). 13C NMR (DMSO-
d6, 100 MHz) (d ppm) 55.6 (CH2), 127.2 (2CH), 127.3 (4CH), 127.9 (2CH), 132.0
(2C), 137.8 (2C), 169.9 (C). 1H NMR (CDCl3, 400 MHz) (d ppm) 3.10–3.40 (br s,
3H), 3.60–3.64 (br s, 2H), 7.26 (t, J = 7.6 Hz, 2H), 7.34 (t, J = 7.6 Hz, 2H), 7.46 (d,
J = 7.6 Hz, 2H), 7.51 (d, J = 7.6 Hz, 2H). 13C NMR (CDCl3, 100 MHz) (d ppm) 55.8
(CH2), 126.9 (2CH), 127.2 (4CH), 128.1 (2CH), 133.1 (2C), 137.7 (2C), 170.6 (C).
MS ESI m/z: 271 (M+, 3), 199(100), 198 (75), 167 (25), 211 (15), 241 (13).
27. Ethyl
(2E)-3-{(2E)-2-[2-oxo-2-(10H-phenothiazin-10-yl)ethylidene]hydrazino}
34. 10-[(5-Ethoxy-3-methyl-1H-pyrazol-1-yl)acetyl]-10H-phenothiazine (17). By-
product from the synthesis of pyrazolone 4a; white solid, 62% yield; mp
but-2-enoate (13).
A
mixture of hydrazone (0.5 mmol) and ethyl
7
141–144 °C. IR (ATR) (m H
max/cmÀ1): 1707, 1559, 1459, 1362, 1258, 1177, 750. 1
acetoacetate 11 (1.0 mmol) in DMA (2 mL) was stirred at 80 °C for 24 h. After
cooling to rt, the mixture was washed with distilled H2O. The resulting yellow
precipitate was filtered. The solid was then purified by column
chromatography on silica gel using EtOAc/n-hexane 1/1 to afford the pure
NMR (CDCl3, 400 MHz) (d ppm) 1.33 (t, J = 7.2 Hz, 3H), 2.18 (s, 3H), 4.02 (q,
J = 7.2 Hz, 2H), 4.80 (br s, 2H), 5.29 (s, 1H), 7.24 (t, J = 7.6 Hz, 2H), 7.32 (dt,
J = 7.6, 1.2 Hz, 2H), 7.44 (d, J = 7.6 Hz, 2H), 7.59 (d, J = 7.6 Hz, 2H). 13C NMR
(CDCl3, 100 MHz) (d ppm) 14.6 (2CH3), 48.9 (CH2), 67.6 (CH2), 84.8 (CH), 127.0
(2CH), 127.1 (2CH), 127.2 (2CH), 128.0 (2CH), 133.1 (2C), 137.8 (2C), 148.2 (C),
155.7 (C), 166.1(C). MS ESI m/z (rel%): 365 (M+, 25), 139 (100), 198 (38), 111
(34), 199 (27).
compound 13 as a yellow solid in 63% yield; mp 152–155 °C. IR (ATR) (mmax
/
cmÀ1): 1677, 1636, 1566, 1460, 1256, 1123, 770, 759. 1H NMR (CDCl3,
400 MHz) (d ppm) 1.25 (t, J = 7.2 Hz, 3H), 2.11 (s, 3H), 4.08 (q, J = 7.2 Hz, 2H),
4.80 (s, 1H), 7.24 (t, J = 7.6 Hz, 2H), 7.29 (s, 1H), 7.32 (t, J = 7.6 Hz, 2H), 7.44 (d,
J = 7.6 Hz, 2H), 7.58 (d, J = 7.6 Hz, 2H). 13C NMR (CDCl3, 100 MHz) (d ppm) 14.3
(CH3), 18.7 (CH3), 59.5 (CH2), 89.2 (CH), 126.7 (2CH), 127.0 (2CH), 127.2 (2CH),
127.9 (2CH), 131.9 (CH), 132.3 (2C) 138.0 (2C), 157.7 (C), 160.9 (C), 169.7 (C).
MS ESI m/z: 381.1 (M+, 17), 199 (100), 198 (89), 167 (15).
35. (a) Chen, X.; She, N.; Shang, Z.; Wu, J.; Wu, H.; Zhang, P. Synthesis 2008, 3478–
3486; (b) Krohn, K.; Stenns, C. Arch. Pharm. 1989, 322, 351–354.
36. 2-[2-Oxo-2-(10H-phenothiazin-10-yl)ethyl]-5-phenyl-2,4-dihydro-3H-pyrazol-3-
one (4b). Yellow solid; 73% yield; mp 187–189 °C. IR (ATR) (m
max/cmÀ1): 1690,
1567, 1458, 1369, 1257, 749. Ketone form: 1H NMR (CDCl3, 400 MHz) (d ppm)
3.62 (s, 2H), 4.71 (br s, 2H), 7.25 (t, J = 7.6 Hz, 2H), 7.34 (t, J = 7.6 Hz, 2H), 7.35–
7.40 (m, 3H), 7.45 (d, J = 7.6 Hz, 2H), 7.58–7.64 (m, 4H). 13C NMR (CDCl3,
100 MHz) (d ppm) 37.6 (CH2), 47.0 (CH2), 125.8 (2CH), 126.8 (2CH), 127.2
(2CH), 127.3 (2CH), 128.1 (2CH), 128.7 (2CH), 130.3 (CH), 130.9 (C), 133.1 (2C),
137.6 (2C), 155.6 (C), 165.5 (C), 172.7 (C). MS ESI m/z: 399.1 (M+, 17), 199 (100),
173 (52), 198 (27).
28. Kamogawa, H.; Larkin, J. M.; Toꢀei, K.; Cassidy, H. G. J. Polym. Sci. Part A 1964, 2,
3603–3614.
29. 10-Acetyl-10H-phenothiazine (15). The hydrazine 6 (10 mmol) was stirred in
absolute EtOH at reflux for 24 h. A white precipitate formed during the reaction
progress and, after cooling to room temperature, it was collected by filtration
and washed with ethanol to afford pure compound 15 as a white solid with the
same physico-chemical properties as described in the literature;28 90% yield.
1H NMR (CDCl3, 400 MHz) d 2.19 (s, 3H), 7.19 (dt, J = 7.6, 1.2 Hz, 2H), 7.30 (dt,
J = 7.6, 1.2 Hz, 2H), 7.41 (dd, J = 7.6, 1.2 Hz, 2H), 7.49 (d, J = 7.6 Hz, 2H).
30. (a) Kennedy, D. A.; Vembu, N.; Fronczek, F. R.; Devocelle, M. J. Org. Chem. 2011,
76, 9641–9647; (b) Morrison, R. W., Jr.; Styles, V. L. J. Org. Chem. 1982, 47, 674–
680; (c) Atkinson, R. S.; Kelly, B. J.; Williams, J. J. Chem. Soc., Chem. Commun.
1992, 373–374; (d) Rigo, B.; Akué-Gédu, R. Targets Heterocycl. Syst. 2006, 10,
232–265. and references cited therein; (e) Akué-Gédu, R.; Couturier, D.;
Hénichart, J.-P.; Rigo, B.; Sanz, G.; Van Hijfte, L.; Bourry, A. Tetrahedron 2012,
68, 1117–1127; (f) Akué-Gédu, R.; Couturier, D.; Hénichart, J.-P.; Rigo, B.; Sanz,
G.; Van Hijfte, L.; Bourry, A. Tetrahedron 2012, 68, 5644–5654; (g) Ghinet, A.;
Farce, A.; Oudir, S.; Pommery, J.; Vamecq, J.; Hénichart, J.-P.; Rigo, B.; Gautret, P.
Med. Chem. 2012, 8, 942–946.
37. General one-pot procedure for the synthesis of pyrazolones: Hydrazine hydrate
(5 mmol) was stirred with 4 Å molecular sieves (0.4 g) in DMA (2 mL) and
cooled to 0–5 °C. A solution of halogenated derivatives 5, 19, or 20 (2.5 mmol)
in DMA (3 mL) was then added. The mixture was stirred at 5–10 °C until
complete consumption of the halogenated reagent (reaction step monitored by
TLC). Ethyl acetoacetate (11) (5 mmol) or ethyl benzoylacetate (16) (5 mmol)
was added and the mixture was stirred at rt for 3 h and then at 60 °C for 10 h.
After cooling to rt, the suspension was filtered in order to remove the
molecular sieves. A mixture of ice/distilled H2O was added to the filtrate. The
resulting solid was collected by filtration and further purified by column
chromatography on silica gel using CHCl3/EtOH 8/2 as eluent to afford pure
pyrazolones 4a–d.
31. Standard procedure for the synthesis of pyrazolones 4a and 4b: A mixture of
hydrazine 6 (5.5 mmol) and ethyl acetoacetate (11) or ethyl benzoylacetate
(16) (5.5 mmol) in DMA was heated at 110 °C for 24 h. After cooling to room
temperature, the mixture was concentrated in vacuo. The crude product was
purified by column chromatography on silica gel using EtOAC/n-hexane 1/1 as
eluent to afford pure pyrazolones 4a and 4b.
38. Cell proliferation assay: Pyrazolone 4b was tested against a panel of 60-human
cancer cell lines at the National Cancer Institute, Bethesda, MD. The
cytotoxicity studies were conducted using a 48 h exposure protocol using
the sulforhodamine B assay. (a) Boyd, R. B. The NCI in vitro Anticancer Drug
Discovery Screen. In Anticancer Drug Development Guide; Preclinical Screening,
Clinical Trials, and Approval; Teicher, B., Ed.; Humana Press Inc. Totowa: NJ,
1997; pp 23–42; (b) Skehan, P.; Storeng, R.; Scudiero, D.; Monks, A.; McMahon,
J.; Vistica, D.; Warren, J. T.; Bokesh, H.; Kennedy, S.; Boyd, M. R. J. Natl. Cancer
Inst. 1990, 82, 1107.
32. Kidwai, M.; Jain, A.; Poddar, R. J. Organomet. Chem. 2011, 696, 1939–1944.
33. 5-Methyl-2-[2-oxo-2-(10H-phenothiazin-10-yl)ethyl]-2,4-dihydro-3H-pyrazol-3-
one (4a): yellow solid; 14% yield; mp 229–231 °C. IR (ATR) (
m
max/cmÀ1): 1689,
1561, 1459, 1260, 1026, 761.
N
N
OH
O
N
N
O
O
N
S
N
S
4a
in CDCl3
4a'
in DMSO-d6