Synthesis and antibacterial activity of some new heterocycles incorporating phthalazine

Article abstract of DOI:10.1016/j.ejmech.2009.06.003

3-(1,4-Dioxo-3,4,4e,5,10,10a-hexahydro-1H-5,10-benzeno-benzo[g]phthalazin-2-yl)-3-oxo-propiononitrile (1) was utilized as key intermediate for the synthesis of some new iminocoumarin 2, chromenone 3, aminothiazole 4, triazepine 5a, b and 6, hydrazono-prop

Full text of DOI:10.1016/j.ejmech.2009.06.003

European Journal of Medicinal Chemistry 44 (2009) 4448–4454
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and antibacterial activity of some new heterocycles
incorporating phthalazine
*
A.M. Khalil, M.A. Berghot, M.A. Gouda
Chemistry Department, Faculty of Science, Mansoura University, Mansoura, 35516, Egypt
a r t i c l e i n f o
a b s t r a c t
Article history:
3-(1,4-Dioxo-3,4,4e,5,10,10a-hexahydro-1H-5,10-benzeno-benzo[g]phthalazin-2-yl)-3-oxo-propiononi-
trile (1) was utilized as key intermediate for the synthesis of some new iminocoumarin 2, chromenone 3,
aminothiazole 4, triazepine 5a, b and 6, hydrazono-propiononitrile 7, pyridopyrazotriazine 8, mono-
bromo 9, dibromo 10 quinoxaline 11, ketene N,S-acetal 13, ketene S,S-diacetal 17 and 18a, b and methyl
dithioate 20 derivatives, respectively. The newly synthesized compounds were characterized by IR, 1H
NMR, 13C NMR and mass spectral studies. Representative compounds of the synthesized product were
tested and evaluated as antibacterial agent.
Received 31 October 2008
Received in revised form
30 May 2009
Accepted 4 June 2009
Available online 13 June 2009
Keywords:
Phthalazine
Ó 2009 Elsevier Masson SAS. All rights reserved.
Coumarine
Triazine
Triazepine
Ketene N,S-acetal
Ketene S,S-diacetal
Bacillus thuringiensis
Escherichia coli
1. Introduction
according to the previous reported method [7]. Cyanoacetic acid
hydrazide was reacted with dibenzobarallene in dimethylformamide
In the past decades, the synthesis of heterocyclic compounds
has been a subject of great interest due to their wide applicability.
Heterocyclic compounds occur very widely in natural and are
essential to life. Among a large variety of heterocyclic compounds,
heterocycles containing phthalazine moiety are of interest because
the show some pharmacological and biological activities (Fig. 1)
[1–3]. Phthalazine derivatives were reported to possess anticon-
vulsant [4], cardiotonic [5], and vasorelaxant [6], activities.
In view of the above mentioned findings and as continuation of
our effort [7–10], to identify new candidates that may be of value in
designing new, potent, selective and less toxic antimicrobial agent,
we report herein the synthesis of some new heterocycles incor-
porating phthalazine moiety starting from dibenzobarallene.
to afford 3-(1,4-dioxo-3,4,4e,5,10,10a-hexahydro-1H-5,10-benzeno-
benzo[g]phthalazin-2-yl)-3-oxo-propiononitrile (1). The structure 1
was established on the basis of elemental and spectral data. The IR
spectrum showed bands at 3200, 2250, 1727 and 1658 cmꢀ1 due to
NH, CN and 3CO groups, respectively. Its 1H NMR spectrum displayed
singlet signals at d 3.7 and 9.8 ppm due totwo protons at CH2and one
proton of NH, respectively. Moreover, the mass spectrum of 1
exhibited the molecular ion peak at m/z 357, which is in agreement
with its molecular formula C21H15N3O3.
Thus, cyclocondensation of compound 1 with salicaldehyde in
boiling dimethylformamide containing a catalytic amount of trie-
thylamine afforded the iminocoumarin derivative 2, which hydro-
lysis in a mixture of Conc. hydrochloric acid and ethanol to achieve
2-chromone derivative 3.
2. Results and discussion
The reaction of
1 with elemental sulfur and phenyl
isothiocyanate in warming dimethylformamide containing a cata-
lytic amount of triethylamine give the thiazole derivative 4.
Structure of the latter product based on analytical and spectral data.
The IR spectrum showed bands at 3414, 3287, 3172 cmꢀ1 (NH) and
1731, 1698 cmꢀ1 (CO). Its 1H NMR spectrum revealed the appear-
2.1. Chemistry
The syntheticprocedures adoptedto obtain the targetcompounds
are depicted in Schemes 1–3. Dibenzobarallene was prepared
ance of signals at
d 6.7 (s, 2H, NH2), 7.1–7.5 (m, 13H, Ar–H) and 9.9
* Corresponding author. Tel.: þ2050 6432235; fax: þ2050 2246781.
E-mail address: dr_mostafa_chem@yahoo.com (M.A. Gouda).
(br, 1H, NH). Moreover, the 13C NMR spectrum characterized by the
0223-5234/$ – see front matter Ó 2009 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2009.06.003

A.M. Khalil et al. / European Journal of Medicinal Chemistry 44 (2009) 4448–4454
4449
CONMe2
of 8 showed three absorption bands at 3405, 3321 and 3271 due to
the three (NH) groups besides three carbonyl absorption bands at
1731, 1692 and 1663.
O
O
O
O
O
CH3
H
AcHN
N
N
N
N
N
N
H
H
Also, its mass spectrum exhibited the molecular ion peak at 530
which is in agreement with the molecular formula C29H22N8O3, the
molecular ion was undergo retro Diels–Alder reaction to revealing
(Mþ-anthracene) and anthracene peaks at m/z 352 and 178.
O
O
HN
NHAr
O
antihypoxic and antipyretic agent [1],
HAV 3C inhibitor [2],
The remarkable importance of a-bromoacetonitrile derivatives
Fig. 1. (a) Antihypoxic and antipyretic agent. (b) HAV 3C inhibitor.
[14,15], for the construction of variety of heterocyclic compounds
promoted us to investigate the synthesis of new bromopropioni-
trile derivative. Thus, a suspension of 1 in acetic acid or chloroform
reacted readily with bromine to yield dibromopropionitrile deriv-
ative 9, while monobromopropionitrile derivative 10 was not
obtained, but when the reaction take place with N-bromosuccina-
mide in dimethylformamide at room temperature compound 9 was
obtained. The structures 9 and 10 were assigned on the basis of
their spectral data. The mass spectrum of compound 10 exhibited
the molecular ion peak at m/z 437 (Mþ þ 2), which is in agreement
with the molecular formula C21H14BrN3O2. Moreover, the 1H NMR
spectrum of compound 9 revealed beside the absence of CH2 signal,
signals of the remaining protons at the expected chemical shift.
Condensation of compound 10 with o-phenylenediamine in
dimethylformamide and in the presence of triethylamine afforded
the aminoquinoxaline derivative 11. The chemical structure of 11
was elucidated on the basis of elemental analysis and spectral data.
The IR spectrum indicated the presence of absorption bands at
3433, 3385 (NH), 1730, 1685 (CO) and 1614 cmꢀ1 (C]N).
presence of three new signals at d 186.1, 152.8 and 79.3 due to C2, C4
and C5 of thiazole ring, respectively.
Cyclization of 4 by refluxing with either triethylorthoformate or
acetic anhydride was studied in the aim of formation of 1,2,4-tri-
azepine derivatives with potential biological activities [11,12]. Thus,
it reacted with either triethylorthoformate or acetic anhydride to
give 1,2,4-triazepine derivatives 5a, b.
The analytical and spectral data are in agreement with the
proposed structure. Thus, IR spectrum of 5a, b showed absorption
bands due to carbonyl groups at 1728–1718 cmꢀ1 beside, the absence
of (NH) and (NH2) bands. The 1H NMR of 5a showed beside the
expected signals, a characteristic signals at 7.1–7.8 (m, 14H, Ar–H,
CH ¼ N). Furthermore, the FAB mass spectrum of 5b showed its quasi-
molecular ion peak appeared at m/z 549 (Mþ þ 1). When compound
5b was heated with dimethylsulphate in dimethylformamide fol-
lowed by stirring with hydrazine hydrate, the hydrazone derivative 6
was formed. Elemental analysis, IR, 1H NMR, 13C NMR and FAB mass
are in agreement with the proposed structure.
Treatment of compound 1 with phenyl isothiocyanate in
dimethylformamide and in the presence of potassium hydroxide, at
room temperature give the non-isolated sodium salt which meth-
ylated by treatment with methyl iodide to afford the novel ketene
N,S-acetal 13. The structure of 13 was established on the basis of its
elemental analysis and spectral data. Its IR spectrum showed
absorption bands at 3286, 3152, 2205 and 1728, 1684 cmꢀ1 due to
two (NH) groups, nitrile and carbonyl functions. The 1H NMR
We have found that diazotized aromatic amine is an excellent
building block for the synthesis of the target compound. Thus,
coupling of compound 1 with 4,6-dimethyl-1H-pyrazolo[3,4-b]pyr-
idine-3-diazonium chloride [13], in pyridine at 0–5 ꢁC afforded the
corresponding hydrazono compound 7. When compound 7 is
refluxed in acetic acid, it can be cyclized tothe corresponding triazine
derivative 8. The formation of 8 may be interpreted through the
nucleophilic attack of ring nitrogen on cyano group. The IR spectrum
O
O
O
H2
C
H
H2N
N
C
CN
CN
O
Phth
DMF
1
O
CHO
OH
TEA / DMF
Dibenzobarallene
H2N
N
O
O
HN
O
O
HCl / EtOH
Ph
PhNCS
Phth
Phth
Phth
S, TEA
S
S
O
O
2
3
4
O
O
Ac2O
or
CH(OEt)3
Phth=
NH
N
R
N
O
R
O
O
Ph
S
Ph
N
N
1- (CH3)2SO4 / TEA / DMF
N
N
N
N
N
N
2- NH2NH2.H2O
MeOH
S
O
O
S
O
NH2
5a, R= H
5b, R= CH3
6
Scheme 1.

4450
A.M. Khalil et al. / European Journal of Medicinal Chemistry 44 (2009) 4448–4454
NH2
spectrum displayed beside the expected signals, a characteristic
singlet signal at 2.1 for three methyl protons. Reaction of 13 with
O
H2N
Phth
N
N
d
CN
NH2
Phth
hydrazine in dimethylformamide not afforded 3-aminopyrazole
14a as expected, but the known phthalazinedione derivative 15
[16], was achieved. Compound 15 was confirmed by its mass
spectrum and comparison of its melting point and mixed melting
point with authentic sample.
Oxoketene dithioacetals, especially dimethyl thioacetal have
considerable attention due to their synthetic importance for the
construction of variety of alicyclic, aromatic and heterocyclic
compounds [17,18]. Thus, propiononitrile derivative 1 treated with
carbon disulphide in the presence of two equivalent sodium
hydride to give the non-isolated sodium salt 16 which was allowed
to react with halogenated compounds namely, methyl iodide,
1,2-dibromomethane or 1,3-dibromopropane to give the corre-
sponding ketene dithioacetal 17 and 18a, b, respectively. IR, 1H NMR
and mass spectra have characterized the structure of these
compounds. The IR spectra exhibited bands at 3440–3300 (NH),
2204–2202 (CN) and 1746–1642 cmꢀ1 (CO). The 1H NMR spectrum
Br
10
TEA / DMF
O
11
O
NBS / DMF
CN
CH3
N2+ Cl-
Phth
HN
CN
CH3
N
N
N
O
H3C
N
N
H
Phth
N
H3C
Pyridine
N
H
1
Br2 / AcOH
or CHCl3
7
O
AcOH
N
CN
Br
Phth
NH2
Br
O
N
CH3
of 17 displayed characteristic two singlet signals at
corresponding to six protons of two methyl groups. Also, 1H NMR
spectrum of 18a showed signals at 3.6 (br, 4H, CH2CH2) and 10.4
d 2.47 and 2.51
9
N
Phth
N
d
N
O
(s, 1H, NH). Furthermore, the (þ)-ESI mass spectrum of 18b showed
four quasi-molecular ion peaks at 474.1 [M þ H]þ, 496.1 [M þ Na]þ,
946.8 [2M þ H]þ and 968.9 [2M þ Na]þ, and (ꢀ)-ESI mass spectrum
showed two quasi-molecular ion peaks at 472.2 [M–H]ꢀ and 944.9
[2 M–H]ꢀ pointing 473 as the molecular mass of the compound.
Refluxing of 13 with hydrazine hydrate not afforded the
aminopyrazole derivative 14 as expected [19], but the phthalazine
derivative 15. While the reaction of compound 1 with carbon
CH3
8
Phth=
NH
N
O
Scheme 2.
O
O
O
CN
CN
CN
H2
C
H2
C
Phth
Phth
Na S
MeI
Phth
Br
(CH2)n
Br
S
S
S Na
SMe
MeS
17
n
16
18a, n= 0
18b, n= 1
CS2 / NaH / DMF
O
O
CN
O
CN
SK
CS2 / NaH
Phth
PhNCS / KOH
DMF
CN
Phth
PhHN
Phth
S
C
C
SNa
1
19
12
MeI
(CH3)2SO4
O
O
O
CN
CN
Phth
CN
Phth
S
Phth
C
C
PhHN
13
SMe
SMe
HS
SCH3
20
NH2NH2.H2O
NH2
O
O
O
O
N
Phth
NH
NH
Phth=
NH
N
NH
R
O
14a, R= NHPh
14b, R= SMe
15
Scheme 3.

A.M. Khalil et al. / European Journal of Medicinal Chemistry 44 (2009) 4448–4454
4451
disulphide and one equivalent sodium hydride in dimethylforma-
mide gave non-isolated sodium dithioacetal derivative 19, which
was alkylated using dimethylsulphate to give methyl dithioate
derivative 20. Compound 20 was confirmed by elemental analysis,
IR, and 1H NMR spectra. The IR spectrum revealed bands at 3136
(NH), 2975 (CH aliphatic), 2202 (CN) and 1730, 1665 (CO). Also, its
Georg-August University Goettingen, Germany and a Brucker AC
300 Eberhard-Karls University, Tuebingen, Germany, in CDCl3 or
DMSO solvent using TMS as internal standard. 13C NMR spectra
were determined on Brucker AC 300 Eberhard-Karls University,
Tuebingen, Germany, in CDCl3 or DMSO solvent using TMS as
internal standard. Mass spectra were recorded on GC–MS QP-1000
EX. Schimadzye (Japan), Faculty of Science, Cairo University, ESI MS
with Quattro Triple Quadruple Mass Finingan MAT–Incos 50 ESILCQ
(Finingan), Georg-August University Goettingen, Germany and
FAB þ Q3MS LMR UP LR, Eberhard-Karls University, Tuebingen,
Germany, Elemental analyses (C, H, and N) were carried out at the
microanalytical center of Cairo Univ., Giza, Egypt, the results were
found to be in good agreement (ꢂ0.12%) with the calculated values.
1H NMR spectrum displayed characteristic signals at
d 3.9 (s, 3H,
CH3), 7.1–7.9 (m, 8H, Ar–H), 10.8 (s, 1H, SH) and 11.12 (s, 1H, NH)
beside the expected signals for other protons.
3. Pharmacology
Thirteen compounds were screened in vitro for their antimi-
crobial activities against Bacillus thuringiensis and Escherichia coli by
the agar diffusion technique [20]. The bacteria were maintained on
nutrient agar. DMSO showed no inhibition zones. The agar media
were incubated with different microorganism culture tested. After
24 h of incubation at 30 ꢁC, the diameter of inhibition zone (mm)
was measured (Table 1). Ampicillin and chloramphenicol were
purchased from Egyptian market and used in a concentration
2 mg mlꢀ1 as references.
The results depicted in Table 1 revealed that compounds 2, 6, 7,
9, 10, 13 and 20 exhibited interesting high activities against the
reference chemotherapeutics. It is worth mentioning that incor-
poration of phthalazine moiety to coumarin, bromo, thiomethyl
and hydrazo group caused significant activity against both B.
thuringiensis and E. coli.
4.1.1. Synthesis of 3-(1,4-dioxo-3,4,4e,5,10,10a-hexahydro-1H-5,10-
benzeno-benzo[g]phthalazin-2-yl)-3-oxo-propiononitrile (1)
A solution of dibenzobarallene (8.28 g, 0.03 mole) and cyano-
acetic acid hydrazide (3.17 g, 0.032 mole) in dimethylformamide
was refluxed for 4–5 h. The separated product was recrystallized
from benzene-dimethylformamide to give 1.
Yield 65%; 7 g; mp 310 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3200 (NH), 2250
(CN),1727, (2CO), and 1658 (CO); 1H NMR (DMSO): dppm ¼ 3.1 (s, 2H,
C11–H, C12–H), 3.7 (s, 2H, CH2–CN), 4.6 (s, 2H, (C9–H, C10–H)),
7.20–7.61 (m, 8H, Ar–H) and 9.8 (s, 1H, NH); EIMS (m/z) (%) ¼ 357
(Mþ, 2.6), 318 (2.9), 290 (4.0), 259 (8.6), 231 (4.0), 202 (8.0), 178
(100), 152 (9.3), 112 (4.0), 82 (5.3) and 55 (3.3). Anal. for C21H15N3O3
(357.36): calcd.: C, 70.58; H, 4.23; N, 11.76%; found: C 70.64, H 4.27,
N 11.83%.
In conclusion, we reported herein a simple and convenient route
for the synthesis of some new heterocyclic based on phthalazine for
antibacterial evaluation.
4.1.2. Synthesis of 2-[(2-imino-2H-chromen-3-yl) carbonyl]-
3,4,4a,5,10,10a-hexahydro-1H-5,10-benzeno-benzo[g]-phthalazine-
1,4-dione (2)
4. Experimental
To a mixture of 1 (1.78; 0.005 mole) and salicyaldehyde (0.61 g;
0.005 mole) in DMF (15 ml) was added TEA (0.3 ml). The reaction
was heated on water bath for 4 h, and poured into ice water. The
obtained product was filtered, washed with water and crystallized
from ethanol-benzene to give 2.
4.1. General
All melting points are in degree centigrade and are uncorrected.
TLC analysis was carried out on silica gel 60 F254 precoated
aluminum sheets Infrared spectra were recorded on FTIR 5300
Yield80%;1.84 g; mp 285 ꢁC;IR (KBr):
n
/cmꢀ1 ¼3353, 3294 (2NH),
1723,1702, (CO),1630(C]N)and1610 cmꢀ1 (C]C);1HNMR(DMSO):
dppm ¼ 3.5 (s, 2H, C11–H, C12–H), 4.8 (s, 2H, (C9–H, C10–H)), 7.0–8.5 (m,
13H, Ar–H), 9.0 (br, 1H, NH (imino)) and 11.8 (s, 1H, NH); EIMS (m/z)
(%) ¼ 461 (Mþ, 4.68), 295 (0.78), 203 (2.42), 202 (3.37), 178 (100), 172
(3.77),102(2.92)and77(3.51). Anal. forC28H19N3O4 (461.47):calcd.:C
72.88, H 4.15, N 9.11%; found: C 72.86, H 4.13, N 9.12%.
spectrometer and Perkin Elmer spectrum RXIFT-IR system (l )
, cmꢀ1
using potassium bromide wafer technique, faculty of science El
Azhar university. 1H NMR spectra were determined on a Varian XL
200 MHz, Faculty of Science, Cairo University, a Brucker WP 300
Table 1
4.1.3. Synthesis of 2-[(2-chromenon-3-yl)carbonyl]-3,4,4a,5,10,10a-
hexahydro-1H-5,10-benzeno-benzo[g]-phthalazine-1,4-dione (3)
Compound 2 (0.92 g; 0.002 mole) was heated in a mixture of
conc. HCl and ethanol (10 ml, 1:1 by vol.) for 15 min. on water bath.
The reaction mixture was left to stand at room temperature over-
night, and the solid product was filtered and crystallized from
ethanol-benzene to give 3.
Inhibition zone (mean diameter of inhibition in mm) as a criterion of antibacterial
activities of the newly synthesized compounds.
Compound no.
Inhibition zone in mm
Gram-positive bacteria
‘‘Bacillus thuringiensis’’
Gram-negative bacteria
‘‘Escherichia coli’’
2
3
4
5a
5b
6
7
9
10
13
18a
18b
20
12
15
11
12
11
13
30
29
20
30
–
20
16
16
13
16
19
20
27
16
26
13
15
22
Yield 83%; 0.76 g; mp > 320 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3256 (NH),
1800, 1726 (CO) and 1610 cmꢀ1 (C]C); 1H NMR (DMSO): dppm ¼ 3.3
(br, 2H, C11–H, C12-H), 4.8 (br, 2H, (C9–H, C10–H)), 7.0–8.6 (m, 13H,
Ar–H), and 10.0 (s, 1H, NH). Anal. for C28H18N2O5 (462.45): calcd.:
C 72.72, H 3.92, N 6.06%; found: C 72.73, H 3.90, N 6.01%.
4.1.4. Synthesis of 2-[(4-amino-3-phenyl-2-thio-(3H)-5-thiazol-5-
yl)carbonyl]-3,4,4a,5,10,10a-hexahydro-1H-5,10-benzeno-benzo[g]-
phthalazine-1,4-dione (4)
To a stirred solution of propiononitrile derivative 1 (3.57 g; 0.01
mole), finally divided sulfur (0.35 g; 0.011 mole) and TEA (1.2 ml) in
a mixture of DMF and EtOH (20 ml, 1:1 by vol.), the phenyl
isothiocyanate (1.35 g; 0.01 mole) was added. The reaction mixture
16
22
Reference drugs
Ampicillin
Chloramphenicol
18
23
19
20

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A.M. Khalil et al. / European Journal of Medicinal Chemistry 44 (2009) 4448–4454
was heated in water bath at 60 ꢁC for 3 h, and then cooled. The
obtained product was filtered, washed with ethanol, dried and
recrystallized from ethyl acetate to give 4.
C12–H), 4.8 (d, 2H, C9–H, C10–H), 6.9–7.4 (m, 10H, Ar–H), 8.8 (br.,
1H, NH–N]C) and 12.2 (s, 1H. NH). Anal. for C29H22N8O3 (530.54):
calcd.: C 65.65, H 4.18, N 21.12%; found: C 65.62, H 4.14, N 21.1%.
Yield 75%, 3.93 g; mp 286 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3414, 3287, 3172
(NH2, NH) and 1731, 1698, 1652 (CO) cmꢀ1
;
1H NMR (DMSO):
4.1.8. Synthesis of pyridopyrazolotriazine derivative 8
7 (1.06 g; 0.002 mole) was refluxed in glacial acetic acid (15 ml)
for 4 h. The reaction mixture was left to stand at room temperature
overnight, and the solid product was filtered and crystallized from
ethanol-benzene to give 8.
dppm ¼ 3.3 (s, 2H, C11–H, C12–H), 4.7 (s, 2H, C9–H, C10–H), 6.7 (s, 2H,
NH2), 7.1–7.5 (m, 13H, Ar–H), and 9.9 (s, 1H, NH). 13C NMR (DMSO):
dppm ¼ 186.1,173.3,169.9,152.8,141.7,138.8,134.4,129.8,126.5,126.0,
124.5, 123.9, 79.3, 44.1 and 44.2. Anal. for C28H20N4O3S2 (524.61):
calcd.: C 64.10, H 3.84, N 10.68%; found: C 64.13, H 3.79, N 10.62%.
Yield 83%, 0.88 g; mp > 320 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3405, 3221,
3271 (NH2, NH), 1731, 1692, 1663 (CO) and 1628 cmꢀ1 (C]N); EIMS
(m/z) (%) ¼ 530 (Mþ, 26.9), 352 (14.8), 241 (12.7), 178 (100), 174
(12.3), 146 (17.5) and 78 (11.8). Anal. for C29H22N8O3 (530.54):
calcd.: C 65.65, H 4.18, N 21.12%; found: C 65.66, H 4.17, N 21.1%.
4.1.5. Synthesis of 6-substituted -4-phenyl-3-thio-1, 8,15-trioxo-
8,8a,9,14,14a,15-hexahydro-3H [10,30] thiazolo [40,50-e]-1H-[1,2,4]
triazepino [1,2-b]-9,14-benzeno-benzo-[g] phthalazine (5a, b)
A solution of compound 4 (1 g; 0.002 mole) in a mixture of
triethylorthoformate and acetic anhydride (15 ml, 3:1 by vol.) or
acetic anhydride was heated under reflux for 3hrs, and then cooled.
The obtained product was filtered, dried and crystallized from
ethanol-benzene to give 5a, b.
4.1.9. Synthesis of 2,2-dibromo-3-(1,4-dioxo-3,4,4a,5,10,10a-
hexahydro-1H-5,10-benzeno-benzo[g]-phthalazin-2-yl)-3-oxo-
propiononitrile (9)
A mixture of bromine (1.6 g; 0.01 mole) in glacial acetic acid or
chloroform (15 ml) was added drop wise over a period of 20 min. to
a hot solution of 1 (3.57 g; 0.01 mole) in acetic acid (75 ml). The
reaction was heated on water bath for 3 hrs; the separated product
after cooling was filtered, washed with water and crystallized from
DMF/MeOH to give 9.
5a; Yield 82%, 0.87 g; mp > 320 ꢁC; IR (KBr):
n
/cmꢀ1 ¼ 2953
(C–H alphatic) and 1718 (br, CO) cmꢀ1; 1H NMR (DMSO): dppm ¼ 3.7
(s, 2H, C11–H, C12–H), 4.9 (s, 2H, C9–H, C10–H) and 7.1–7.5 (m, 14H,
Ar–H, N–CH]N). Anal. for C29H18N4O3S2 (534.61): calcd.: C 65.15, H
3.39, N 10.48%; found: C 65.17, H 3.4, N 10.35%.
5b; Yield 93%, 1.0 g; mp > 320 ꢁC; IR (KBr):
n
/cmꢀ1 ¼ 2950, 2900
Yield 79%, 4.06 g; mp 221 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3250 (NH),
(C–H aliphatic) and 1728 (br, CO) cmꢀ1; FABMS (m/z) (%) ¼ 549,
(Mþ þ 1, 9.60), 460 (4.80), 307 (100), 289 (73), 215 (24.3), 204 (12.1),
202 (16.5),178 (38.7) and 165 (97.8). Anal. forC30H20N4O3S2 (548.63):
calcd.: C 65.68, H 3.67, N 10.21%; found: C 65.69, H 3.63, N 10.20%.
2260 (CN) and 1722, 1662 cmꢀ1(CO). 1H NMR (DMSO): dppm ¼ 3.4
(s, 2H, C11–H, C12–H), 4.8 (s, 2H, (C9–H, C10–H)), 7.1–7.5 (m, 8H,
Ar–H), and 11.6–11.8 (br, 1H, NH). Anal. for C21H13Br2N3O3 (515.15):
calcd.: C 48.96, H 2.54, N 8.16%; found: C 48.86, H 2.50, N 8.20%.
4.1.6. Synthesis of 3-hydazino-6-methyl-4-phenyl-1,8,15-trioxo-
8,8a,9,14,14a,15-hexahydro-3H-[10,30] thiazolo [40,50-e]-1H-[1,2,4]
triazepino [1,2-b]-9,14-benzeno-benzo-[g] phthalazine (6)
4.1.10. Synthesis of 2-bromo-3(1,4-dioxo-3,4,4a,5,10,10a-
hexahydro-1H-5,10-benzeno-benzo[g]-phthalazin-2-yl)-
3-oxo-propiononitrile (10)
To a solution of compound 5b (5.5 g; 0.01 mole) in DMF (20 ml)
dimethylsulphate (1.89 g; 0.015 mol) was added the reaction
mixture was heated on water bath at 90 ꢁC for 3 h. It was then
poured portion wise to a stirred solution of hydrazine hydrate
(2 ml) in methanol (10 ml). The stirring was continued for 2 h. The
obtained product after adding ice cold-water was filtered washed
with EtOH, dried and crystallized form DMF–methanol to give 6.
A mixture of 1 (3.57 g; 0.01 mole) and N-bromosuccinamide
(1.78 g; 0.01 mole) in DMF (30 ml) was stirred for 48 h, at room
temperature. The obtained product after dilution with water was
filtered, washed with water and crystallized from ethanol-benzene
to give 10.
Yield 75%, 3.27 g, mp 306 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3186, 3186 (NH),
2255 (CN) and 1796, 1728 cmꢀ1(CO); EIMS (m/z) (%) ¼ 437 (Mþ þ 2,
0.19), 435 (Mþ, 0.2), 355 (0.11), 313 (0.72), 204 (1.09), 203 (4.39),
202 (6.62) and 178 (100). Anal. for C21H14BrN3O3 (436.26): calcd.:
C 57.82, H 3.23, N, 9.63%; found: C 57.73, H 3.33, N 9.68%.
Yield 78%, 4.25 g; mp 295 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3400, 3336,
3261 (NH, NH2), 1772, 1704, 1682 (CO) and 1642 cmꢀ1 (C]N); 1H
NMR (DMSO): dppm ¼ 2.45 (s, 3H, CH3), 3.2 (s, 2H, NH2), 3.4 (d, 2H,
C11–H, C12–H), 4.8 (d, 2H, C9–H, C10-H) and 7.1–7.6 (m, 13H, Ar–H);
13C NMR (DMSO): dppm ¼ 173.8, 160.0, 154.1, 153.6, 141.8, 139.0,
135.4, 129.5, 129.3, 128.7, 126.4, 126.1, 124.5, 124.1, 44.4, 44.2, 40.5,
40.1 and 21.7; FABMS (m/z) (%) ¼ 547 (Mþ þ 1, 1), 460 (3), 444 (3.5),
307 (33.9), 291 (85.6), 289 (31.3), 215 (7.8), 204 (11.3), 202 (15.6)
and 178 (100). Anal. for C30H22N6O3S (546.60): calcd.: C 65.92,
H 4.06, N 15.38%; found: C 65.89, H 4.02, N 15.39%.
4.1.11. Synthesis of 2-[(3-amino-benzopyridazin-2-yl)carbonyl]-
1,4-dioxo-3,4,4a,5,10,10a-hexahydro-1H-5,10-benzeno-
benzo[g]phthalazine (11)
A mixture of 9 (0.87 g; 0.002 mole), o-phenylenediamine
(0.21 g, 0.02 mole) and triethylamine (3 drops) in DMF (15 ml) was
refluxed for 24 h. The obtained product after dilution with water
was filtered, washed with water and crystallized from ethanol-
benzene to give 11.
4.1.7. Synthesis of 3-(1,4-dioxo-3,4,4a,5,10,10a-hexahydro-1H-5,10-
benzeno-benzo[g]-phthalazin-2-yl)-3-oxo-2-[3-(4,6-dimethyl-1H-
pyrazolo[3,4-b]pyridine)hydrazono]propiononitrile 7
Yield 76%, 0.7 g; mp 273 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3433, 3335 (NH,
NH2), 1730, 1685, (CO) and 1614 cmꢀ1(C ¼ N); EIMS (m/z) (%) ¼ 461
(Mþ,18.3), 290 (1.7), 283 (6.9), 255 (0.7), 204 (1), 203 (4.7), 202 (5.4)
178 (100), 144 (1.7) and 97 (2.5). Anal. for C27H19N5O3 (461.47):
calcd.: C 70.27, H 4.15, N 15.18%; found: C 70.3, H 4.18, N 15.22%.
A
well stirred solution of 3-amino-(4,6-dimethyl-1H-pyr-
azolo[3,4-b]pyridine) (0.362 g; 0.002 mole) in (0.6 ml) concentrated
HCl and H2O (2 ml) was cold in ice-bath and diazotized with the
solution of NaNO2 (0.15 g; 0.002 mole) in H2O (2 ml). The cold
diazonium solution was added slowly to a well stirred solution of 1
(0.70 g; 0.002 mole) in pyridine (10 ml). The reaction mixture was
stirred for another 2 hrs. The crude product was filtered off, dried
well and recrystallized from ethanol-benzene 7.
4.1.12. Synthesis of 2-methylthio-2-phenylaminomethylene-3-
(1,4-dioxo-3,4,4a,5,10,10a-hexahydro-1H-5,10-benzeno-
benzo[g]-phthalazin-2-yl)-3-oxo-propiononitrile (13)
To a cold suspension of finally divided KOH (0.28 g; 0.005 mole)
in dry dimethylformamide (25 ml) were added the nitrile deriva-
tive 1 (1.78 g; 0.005 mole) followed by phenyl isothiocyante. The
mixture was stirred at room temperature for 12 h, then cooled
Yellow crystals; yield 76%; mp > 320; IR (KBr):
n
/cmꢀ1 ¼3382,
3202 (NH), 2235 (CN), 1732, 1686 (CO) and 1508 (N]N); 1H NMR
(DMSO): dppm ¼ 2.6 (s, 3H, CH3), 2.9(s, 3H, CH3–C]N), 3.4 (d, 2H, C11–H,

A.M. Khalil et al. / European Journal of Medicinal Chemistry 44 (2009) 4448–4454
4453
again to 0 ꢁC, treated with the methyl iodide (0.7 g; 0.005 mole)
and left to stand at room temperature for 8 h. The mixture was
poured into ice cold-water. The resulting precipitate was filtered
off, dried and crystallized from DMF-MeOH to give 13.
pointing 473 as the molecular mass of the compound. Anal. for
25H19N3O3S2 (473.57): calcd.: C 63.41, H 4.04, N 8.87%; found:
C 63.52, H 4.00, N 8.82%.
C
Yield 74%, 1.87 g; mp 278 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3286, 3152
4.1.15. Synthesis of methyl-2-cyano-3-(1,4-dioxo-3,4,4a,5,10,10a-
hexahydro-1H-5,10-benzeno-benzo[g]-phthalazin-2-yl)-3-oxo-
propane dithioate (20)
(2NH), 2205(CN) and 1728, 1684, cmꢀ1 (CO); 1H NMR (DMSO):
dppm ¼ 2.1 (s, 3H, CH3), 3.4 (s, 2H, C11–H, C12–H), 4.8 (s, 2H, C9–H,
C10–H), 7.1–7.5 (m, 13H, Ar–H),10.4 (s, 1H, NHPh) and 11.7 (s, 1H,
NH); EIMS (m/z) (%) ¼ 508 (Mþ þ 2, 0.19), 506 (Mþ, 1.55), 432 (0.47),
290 (1.14), 275 (0.29), 231 (0.49), 204 (1.10), 202 (4.44), 178 (100),
97 (0.42) and 77 (57.8). Anal. for C29H22N4O3S (506.57): calcd.:
C 68.76, H 4.38, N 11.06%; found: C 68.63, H 4.30, N 11.00%.
To a cold suspension of NaH (0.12 g; 0.005 mole) in dry dime-
thylformamide (25 ml) were added the nitrile derivative 1 (1.78 g;
0.005 mole) followed by carbon disulphide (1.14 g; 0.15 ml) slowly
drop wise under stirring over a period of 15 min while the
temperature of the mixture was maintained at 5–10 ꢁC. The
mixture was stirred at room temperature for 12 h, then cooled
again to 0 ꢁC, dimethylsulphate (1.5 g; 0.01 mole) was added drop
wise over a period of 10 min. The reaction mixture was stirred for
4 hrs, at room temperature and then poured into ice cold-water.
The resulting precipitate was filtered off, dried and crystallized
from DMF-MeOH to give 20.
4.1.13. Synthesis of 2,3,4,4a,5,10,10a-heptahydro-1H-5,10-benzeno-
benzo[g]-phthalazine-1,4-dione (15)
To suspension of 13 (1.01 g; 0.002 mole) or 17 (2.90 g; 0.002
mole) in DMF/EtOH (10 ml; 1.3 by vol.), hydrazine hydrate (0.1 ml;
0.002 mole) was added. The reaction mixture was heated under
reflux for 2 h. The obtained product after cooling was filtered, dried
and crystallized from the EtOH-benzene to give 15.
Yield 85%, 1.9 g; mp 276 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3136 (NH),
2975(C–H aliphatic), 2202 (CN), and 1730, 1665 cmꢀ1 (CO); 1H NMR
(DMSO): dppm ¼ 3.5 (d, 2H, C11–H, C12–H), 3.9 (s, 3H, CH3), 4.8 (d, 2H,
C9–H, C10–H), 7.1–7.9 (m, 8H, Ar–H), 10.8 (s, 1H, SH) and 11.2 (s, 1H,
NH). Anal. for C23H17N3O3S2 (447.53): calcd.: C 61.73, H 3.83,
N 9.39%; found: C 61.83, H 3.92, N 9.42%.
Yield 81%; mp 290 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3336, 3260 (2NH), and
1774, 1696 cmꢀ1 (CO); EIMS (m/z) (%) ¼ 290 (Mþ, 2.70), 231 (0.2),
204 (0.6), 202 (5.60), 178 (100), 112 (2.40) and 77 (1.70). Anal. for
C18H14N2O2 (290.32): calcd.: C 74.47, H 4.86, N 9.65%; found:
C 74.51, H 4.82, N 9.60%.
4.2. In vitro antimicrobial activity
4.1.14. Synthesis of ketene S,S-acetal (17, 18a, 18b)
The tested compounds were evaluated by the agar diffusion
technique [20] using a 2 mg mlꢀ1 solution in DMSO. The test
organisms were B. thuringiensis (Gram-positive) and E. coli (Gram-
negative). A control using DMSO without the test compound was
included for each organism. Ampicillin and chloramphenicol in
DMSO were used as reference drugs.
4.1.14.1. General procedure. To a cold suspension of NaH (0.24 g;
0.01 mole) in dry dimethylformamide (25 ml) were added the
nitrile derivative 1 (1.78 g; 0.005 mole) followed by carbon disul-
phide (1.14 g; 0.15 ml) slowly drop wise under stirring over a period
of 15 min. while the temperature of the mixture was maintained at
5–10 ꢁC. The mixture was stirred at room temperature for 12 h.
Then cooled again to 0 ꢁC, methyl iodide (1.42 g; 0.01 mole) or
dihalocompound (0.005 mole) namely dibromoethane or dibro-
mopropane was added drop wise over a period of 10 min and left to
stand at room temperature for 24 h. The mixture was poured into
ice cold-water. The resulting precipitate was filtered off, dried and
crystallized from ethanol-benzene.
Acknowledgements
We would to express our thanks to Dr. M. Ismail, Assistant
Professor, Chemistry Department, and Faculty of science, Mansoura
University, Dr. A. Ellabady, Eberhard-Karls University, Tuebingen. K.
Attai Shaaban, Georg-August University, Germany, for spectral
measurements. Also, we would like to express our sincere thanks to
Dr. A. Shawky, Assistant Lecturer, Botany Department, Faculty of
science, Mansoura University, for microbiological screening.
4.1.14.1.1. 2-[Bis(methylthio)-methylene]-3-(1,4-dioxo-
3,4,4a,5,10,10a-hexahydro-1H-5,10-benzeno-benzo[g]-
phthalazin-2-yl)-3-oxo-propiononitrile (17). Yield 75%; 1.72 g, mp
270 ꢁC; IR (KBr):
n
/cmꢀ1 ¼3440 (NH), 2202(CN), 1746, 1694 (CO)
and 1638 cmꢀ1 (C ¼ C); 1H NMR (DMSO): dppm ¼ 2.47 (s, 3H, CH3),
2.51(s, 3H, CH3), 3.4 (s, 2H, C11–H, C12–H), 4.8 (s, 2H, C9–H, C10–H),
7.1–7.5 (m, 8H, Ar–H) and 10.9 (s, 1H, NH); EIMS (m/z) (%) ¼ 461
(Mþ, 1.69), 453 (3.16), 378 (4.24), 313 (8.71), 264 (6.40), 178 (100),
152 (1.23) and 57 (38.44). Anal. for C24H19N3O3S2 (461.56): calcd.:
C 62.45, H 4.15, N 9.10%; found: C 62.59, H 3.96, N 9.1%.
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