Synthesis, characterization, and biological-pharmacological evaluation of new phosphazenes bearing dioxybiphenyl and Schiff base groups

Article abstract of DOI:10.1002/ardp.201100412

A new spirocyclophosphazene, 2,2-bis(2-formylphenoxy)-4,4,6,6- bis[spiro(2′,2″-dioxy-1′-1″-biphenylyl)] -cyclotriphosphazene (3), was obtained from the reaction of 2,2-dichloro-4,4,6, 6-bis[spiro(2′,2″-dioxy-1′-1″-biphenylyl)] cyclotriphosphazene (2) with 2-hydroxybenzaldehyde. New phosphazene derivatives bearing Schiff base and dioxybiphenyl groups have been synthesized by the reactions of 3 with different amines. The structures of the compounds were defined by elemental analysis, IR, ¹H, ¹³C, and ³¹P NMR spectroscopy. All the synthesized compounds were screened for their antibacterial activity against both types of Gram-negative and Gram-positive bacteria. The most potent antibacterial compound of this series was compound 12 which has the low MIC value of 3.75-0.9375 μg/mL. Both minimal inhibitory concentrations (MIC) and inhibition zones were determined in order to monitor the efficacy of the synthesized compounds. New compounds were also screened for anticonvulsant, CNS depressant, and sedative-hypnotic activity. After i.p. injection to mice at doses of 30, 100, and 300 mg/kg body weight phosphazenes were examined in the maximal electroshock-induced seizure (MES) and subcutaneous pentylenetetrazole (scPTZ) induced seizure models in mice. The neurotoxicity was assessed using the rotorod method. Compounds 4, 5, and 11 were found to be active in both MES screen and scPTZ screen at 0.5 h. All except 12 showed more than 44% decrease in locomotor activity after 1 h of compound administration via actophotometer screen. CNS-depressant activity screened with the help of the forced swim method resulted in some potent compounds. Except for 7 and 12 other tested compounds were found to exhibit potent CNS depressants activity as indicated by increased immobility time.

Full text of DOI:10.1002/ardp.201100412

Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
1
Full Paper
Synthesis, Characterization, and Biological–Pharmacological
Evaluation of New Phosphazenes Bearing Dioxybiphenyl and
Schiff Base Groups
Erol Cil1, Mehmet Ali Tanyildizi2, Furkan Ozen2, Mustafa Boybay2, Mustafa Arslan3, and
Ahmet Orhan Gorgulu2
1 Education Faculty, Science Education Department, Firat University, Elazig, Turkey
2 Science Faculty, Chemistry Department, Firat University, Elazig, Turkey
3 Education Faculty, Science Education Department, Yildiz Technical University, Istanbul, Turkey
A new spirocyclophosphazene, 2,2-bis(2-formylphenoxy)-4,4,6,6-bis[spiro(20,200-dioxy-10-100-biphenylyl)]-
cyclotriphosphazene (3), was obtained from the reaction of 2,2-dichloro-4,4,6,6-bis[spiro(20,200-dioxy-10-
100-biphenylyl)]cyclotriphosphazene (2) with 2-hydroxybenzaldehyde. New phosphazene derivatives
bearing Schiff base and dioxybiphenyl groups have been synthesized by the reactions of 3 with
different amines. The structures of the compounds were defined by elemental analysis, IR, 1H, 13C, and
31P NMR spectroscopy. All the synthesized compounds were screened for their antibacterial activity
against both types of Gram-negative and Gram-positive bacteria. The most potent antibacterial
compound of this series was compound 12 which has the low MIC value of 3.75–0.9375 mg/mL.
Both minimal inhibitory concentrations (MIC) and inhibition zones were determined in order to
monitor the efficacy of the synthesized compounds. New compounds were also screened for
anticonvulsant, CNS depressant, and sedative-hypnotic activity. After i.p. injection to mice at
doses of 30, 100, and 300 mg/kg body weight phosphazenes were examined in the maximal
electroshock-induced seizure (MES) and subcutaneous pentylenetetrazole (scPTZ) induced seizure
models in mice. The neurotoxicity was assessed using the rotorod method. Compounds 4, 5, and
11 were found to be active in both MES screen and scPTZ screen at 0.5 h. All except 12 showed more
than 44% decrease in locomotor activity after 1 h of compound administration via actophotometer
screen. CNS-depressant activity screened with the help of the forced swim method resulted in some
potent compounds. Except for 7 and 12 other tested compounds were found to exhibit potent CNS
depressants activity as indicated by increased immobility time.
Keywords: Antibacterial activity / Anticonvulsant / Hexachlorocyclotriphosphazene / Phosphazene
Received: November 22, 2011; Revised: January 16, 2012; Accepted: January 24, 2012
DOI 10.1002/ardp.201100412
Introduction
or as photoactive materials [2–8]. They also possess a number
of characteristics such as biomedical properties and appli-
cations due to their strong antitumor activity [9–13]. Their
antimicrobial and biological activities on bacterial and yeast
cells have been studied [14–16]. Some applications include
model compounds for polyphosphazenes, starting materials
for the preparation of cyclolinear and/or cyclomatrix
phosphazene substrates, commercial polymers with carbon
backbones containing pendant cyclophosphazene groups,
inorganic hydraulic fluids and lubricants, biologically
important substrates such as anticancer agents, insect
chemosterilants, pesticides and fertilizers, supports for cata-
lysts, dyes, and crown ether phase transfer catalysts for
Phosphazenes are compounds that contain a framework of
alternating phosphorus and nitrogen atoms, either in cyclic
or linear form [1]. These compounds possess interesting prop-
erties. For example, they exhibit fire-retardant properties,
have high refractive indices, and might find application in
non-linear optics, as ferroelectric materials, as liquid crystals,
Correspondence: Erol Cil, Education Faculty, Science Education
Department, Firat University, 23169 Elazig, Turkey.
E-mail: cilerol@yahoo.com
Fax: (þ90) 424 2330062
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2
E. Cil et al.
Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
nucleophilic substitution reactions, core substrates for
dendrimers, thermal initiators for anionic polymerization
reactions, and photosensitive materials [17].
of 3 with R-(ꢀ)-1-cyclohexylamine and 4-methylbenzylamine.
All the synthesized compounds have been screened for their
antibacterial activity against both types of Gram-negative
and Gram-positive bacteria. New compounds were also
screened for anticonvulsant, CNS depressant, and sedative-
hypnotic activity. The present work was carried out on three
objectives: (i) The first objective was the synthesis and anti-
microbial evaluation of the new phosphazene compounds;
(ii) the second objective was anticonvulsant screening of the
synthesized phosphazenes by MES and subcutaneous pentyl-
enetetrazole (scPTZ) methods; (iii) the third objective was the
sedative-hypnotic, CNS depressant, and phenobarbitone
induced hypnosis potentiation screening of the synthesized
compounds.
The literature contains reports on the synthesis of different
linear, cyclic, or poly-phosphazenes [18–28]. The synthesis
and different reactions of phosphazenes containing 2,20-
dioxybiphenyl groups are reported [29, 30]. There are also
a large number of literature reports on reactions of the
functional groups on phosphazene substituents [31].
Typical of these include coupling reactions of trimeric phos-
phazene azides with aryloxy, alkoxy, and dialkylamino
cosubstituents [32], N-vinylic phosphazenes with azodicar-
boxylic and acetylenic esters [33], oxime-phosphazene deriva-
tives with alkyl and acyl substituents [34–37], polymers
from 4-formylphenoxy [38, 39], maleic [40], and 3,4-methyl-
enedioxyphenoxy substituents [41].
Results and discussion
Cyclophosphazenes containing Schiff bases have been iso-
lated by three methods. The reaction of aryloxycyclophos-
phazenes containing aldehyde group with primary amines,
the reaction of halocyclophosphazenes with Schiff bases
bearing hydroxy, and the reaction of aldehydes with amine
bonded aryloxycyclophosphazenes (achieved by the catalytic
reduction of phosphazene containing nitrophenoxy). Schiff
bases have a variety of applications in fields such as flame
resistance, biomedicine, semi-conductors, dyes, and catalysis
[42]. Antiproliferative activity of phosphazenes containing
Schiff bases was observed to be much higher against cancer
cells than the normal cells [42].
The reaction of 2 with 2 equiv. of 2-hydroxybenzaldehyde in
the presence of K2CO3 in THF gave 2,2-bis(2-formylphenoxy)-
4,4,6,6-bis[spiro(20,200-dioxy-10-100-biphenylyl)] cyclotriphos-
phazene (3). Disubstituted compounds bearing Schiff bases
were obtained from the reactions of 3 with 4-methoxybenzyl-
amine, 2-chlorobenzylamine, 4-chlorobenzylamine, 2-naph-
thylethylamine, 2-aminophenol, 4-aminophenol, biphenyl-2-
amine, and 2-amino-4-chlorophenol. However, monosubsti-
tuted products were obtained from the reactions of 3 with
4-aminobenzonitrile and pyrimidine-2-amine. Pure and
defined Schiff base products could not be obtained from
the reactions of 3 with R-(ꢀ)-1-cyclohexylamine and 4-meth-
ylbenzylamine. All products were generally obtained in high
yields. The structures of the compounds were defined by
elemental analysis, IR, 1H, 13C, and 31P NMR spectroscopy.
The synthetic route of the title compounds is outlined in
Scheme 1 and the structures of the compounds 2–13 are
shown in Scheme 2.
Epilepsy is one of the most common diseases of the brain
affecting at least 50 million persons worldwide. There is
currently a need for improved agents for the treatment of
seizures. Polytherapy with antiepileptic drugs is necessary in
clinical practice because of the limited efficacy of monother-
apy. In recent years many new chemical entities have been
designed that were structurally dissimilar from many com-
mon anticonvulsants containing dicarboximide function,
which contributes to toxic side effects. However, the litera-
ture does not contain any report on the synthesis, antibacte-
rial, and anticonvulsant activity of the cyclophosphazenes
containing dioxybiphenyl and Schiff base groups. In the
present study, we synthesized new spirocyclophosphazene,
2,2-bis(2-formylphenoxy)-4,4,6,6-bis[spiro(20,200-dioxy-10-100-
biphenylyl)]cyclotriphosphazene (3) from the reaction of 2,2-
dichloro-4,4,6,6-bis[spiro(20,200-dioxy-10-100-biphenylyl)]cyclo-
triphosphazene (2) with 2-hydroxy-benzaldehyde. Spirocyclo-
phosphazenes containing Schiff bases were hereby isolated
for the first time from the reactions of 3 with amines such as
4-methoxybenzylamine, 2-chlorobenzylamine, 4-chloroben-
zylamine, 2-naphthylethylamine, 2-aminophenol, 4-amino-
phenol, 4-aminobenzonitrile, biphenyl-2-amine, 2-amino-4-
chlorophenol, and pyrimidine-2-amine. Pure and defined
Schiff base products could not be obtained from the reactions
The characteristic stretching peaks in the IR spectra of the
phosphazenes have been assigned as in experimental section.
–
The P N stretching vibrations, which were observed between
–
1166 and 1194 cmꢀ1, are characteristic of cyclophospha-
zenes. Compared to 1, which appeared at 1218 cmꢀ1, these
peaks are shifted to longer wavelengths for 2–13. In the IR
–
spectra of 4–13, the C N stretching vibrations which were
–
observed between 1619 and 1659 cmꢀ1 indicate the Schiff
bases. The carbonyl stretching vibrations in the IR spectra of
3, 10, and 13 indicate the aldehyde group. None of the aldehyde
–
groups in 3 could be converted into –C N– groups in 10 and
–
13 by the 4-aminobenzonitrile and pyrimidine-2-amine sub-
stituents, respectively. The bands for phenolic OH group for
8, 9, and 12 were observed at 3422, 3341, and 3370 cmꢀ1
,
respectively.
The NMR data of compounds 2–13 were given in exper-
imental section. An example spectrum (31P NMR spectrum of
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Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
Synthesis, Characterization, and Biological–Pharmacological Evaluation of New Phosphazenes
3
Cl Cl
P
N
N
Cl Cl
P
O
O
P
P
O
N
P
N
N
O
+
Cl
P
Cl
N
Cl
Cl
HO
OH
O
( )
1
(2)
HO
O
R
N
O
O
N
R
R
N
O
O
O
O
O
O
P
N
R - NH2
R - NH2
P
N
P
N
N
N
N
N
N
O
O
P
P
O
O
O
O
P
P
O
O
P
P
O
O
N
O
O
(
10,13
)
3
( )
(
4,5,6,7,8,9,11,12)
Scheme 1. Synthetic route of phosphazene derivatives.
13
20
OH
O
7
O
13
10
4
2
5
4
2
5
O
7
N
4
2
12
11
5
6
6
12
3
14
11 15
19
18
6
N3P3
N3P3
3
8
3
N3P3CI2
8
1
10
9
1
1
O
O
O
O
O
O
16
17
9
2
2
2
2
2
(2)
3
( )
8
( )
13
12
11
13
12
11
4
2
4
2
5
O
7
N
5
O
7
N
6
6
O
14
N3P3
3
3
18
17
14 15
N3P3
CH3
19
8
8
15
16
1
1
18
OH
17
O
O
O
O
10
10
16
9
9
2
2
2
2
(
9
)
( )
4
13
12
11
24
4
Cl
5
O
N
O
O
19
13
12
11
6
23
22
14
7
18
19
4
2
20
5
O
N
N3P3
3
8
15
6
7
18
1
2
3
14 15
17
N3P3
N3P3
N3P3
O
O
10
10
16
9
21
20
CN
25
8
16 17
2
1
(
)
O
O
10
9
23
2
2
22
(
5
)
4
5
13
O
7
N
21
6
20
16
15
12
14
1119
N3P3
N3P3
N3P3
3
13
12
11
8
4
2
5
O
7
N
1
2
O
O
6
Cl
10
17
9
18
3
18
17
14 15
2
2
8
(
11
)
16
22
1
13
12
11
O
O
10
9
O
7
N
OH
15
4
5
2
2
14
6
6
( )
3
16
8
19
1
2
10
17
9
21
19
18
O
O
Cl
13
12
11
23
24
15
4
2
2
5
O
7
N
20
(
12
)
6
14
3
18
17
8
13
23
H3C
16
4
1
2
5
O
7
N
O
O
25
O
O
10
9
N
6
12
14
N
17
18
22
21
2
3
2
8
11
7
( )
1
2
16
O
O
15
10
9
20
19
2
(
13
)
Scheme 2. Structures of compounds 2–13.
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4
E. Cil et al.
Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
Figure 1. The 31P NMR spectrum of 9.
9) is presented in Fig. 1. In the 31P NMR spectra of 2–13, two
signals were observed as one doublet and one triplet (AX2
system). The 31P NMR shifts of 2–13 change between 9.39 and
26.60 ppm. This data demonstrates that compounds 2–13
have one isomer. However, in our similar published studies,
we also observed weak peaks due to the syn and anti isomer-
The antibacterial activities of all the synthesized com-
pounds (2–13) and the standard drugs ciprofloxacin and
gatifloxacin were assessed against both types of Gram-nega-
tive bacteria (Pseudomonas aeruginosa ATCC 27853, Klebsiella
pneumoniae CIP 53153, and Escherichia coli ATCC 25922) and
Gram-positive bacteria (Staphylococcus aureus ATCC 25923).
Diameter of zone of inhibition in mm and minimal inhibi-
tory concentration (MIC) in mg/mL of all the compounds are
given in Tables 1 and 2. Dimethyl sulfoxide treated group
served as a control. The compounds 2–13 exhibited varying
antibacterial response against different types of bacterial
strains, but the compounds having groups such as OH, Cl,
and CN showed better results than the other compounds. The
compounds 6, 7, and 11 were devoid of antibacterial activity
against Gram-negative bacteria P. aeruginosa ATCC 27853 and
K. pneumoniae CIP 53153. These compounds showed mild
antibacterial activity against E. coli ATCC 25922 with MIC
of 120, 250, and 120 mg/mL, respectively. The compounds 6
and 7 exhibited moderate activity against Gram-positive bac-
teria S. aureus ATCC 25923 with MIC of 60 and 90 mg/mL,
respectively. Among the compounds 2–11, compound 10
having CN and CO groups exhibited good antibacterial
activity against Gram-negative bacteria P. aeruginosa ATCC
27853, E. coli ATCC 25922, and Gram-positive bacteria S. aureus
ATCC 25923 with MIC 15 mg/mL for each. Moreover, com-
pound 9 showed good activity against Gram-negative bacteria
E. coli ATCC 25922 with MIC 15 mg/mL and moderate anti-
bacterial activity against K. pneumoniae CIP 53153 with MIC
30 mg/mL. Furthermore, the compound 12 (having OH and Cl)
has shown most potent antibacterial activity against Gram-
negative bacteria P. aeruginosa ATCC 27853 (MIC 3.75 mg/mL),
K. pneumoniae CIP 53153 (MIC 1.875 mg/mL), E. coli ATCC 25922
(MIC 3.75 mg/mL). Compound 12 exhibited maximum anti-
–
ism of the –C N groups, so we obtained compounds that are
–
mixtures of syn and anti isomers from the reactions of hex-
akis(4-[(hydroxyimino)methyl]phenoxy) cyclotriphosphazene
and hexakis(4-[(1)-N-hydroxyethanimidoyl]phenoxy)cyclotri-
phosphazene with different alkyl and acyl halogens [34,
35]. It may be that minor isomers were removed during
purification, so we observed only one isomer in this study.
The 1H and 13C NMR data also confirm the structures of 2–
13 (Scheme 1). The detailed 1H NMR spectral data were given
in experimental section. In the 1H NMR spectra, the aldehyde
protons were observed at 10.44 ppm for 3, 10, and 10.43 ppm
for 13. The imine protons for 4–13 were observed between 8.06
and 8.85 ppm. It is understood from the integral intensities
that there are two aldehyde protons in 3, and one aldehyde
proton in 10 and 13. The integral intensities also show that
there is one imine proton in 10 and 13, and two imine protons
in 4–9, 11, and 12. These observations for 10 and 13 indicate
that only one aldehyde group was converted into imine group
in 10 and 13. The phenolic OH protons for 8, 9, and 12 were
observed at 9.09, 8.77, and 10.50 ppm, respectively.
The detailed 13C NMR spectral data were given in exper-
imental section. All of the carbon peaks in the compounds
are visible in the 13C NMR spectra, as expected. The aldehyde
carbon atoms were observed at 188.69 ppm for 3 and 10 and
188.72 ppm for 13. The imine carbon atoms in the substi-
tuted moiety of compounds 4–13 were observed between
152.37 and 166.73 ppm.
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Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
Synthesis, Characterization, and Biological–Pharmacological Evaluation of New Phosphazenes
5
Table 1. Inhibitory-zone diameter (in mm) of the synthesized compounds 2–13 against the tested bacterial strains
Comp. no.
Diameter of the inhibition zone (in mm) Antibacterial activity
LD50 mg/kg
Pseudomanas
aeruginosa
ATCC 27853
Klebsiella
pneumoniae
CIP 53153
Escherichia
coli
ATCC 25922
Staphylococcus
aureus
ATCC 25923
2
3
4
5
6
7
8
9
10
–
8
11
13
–
7
–
16
11
–
10
8
–
18
9
8
11
22
22
11
27
13
23
26
–
–
>800
>800
>800
>800
>800
>800
>800
>800
>800
>800
>1600
>800
12
14
16
13
11
13
21
–
29
11
26
24
–
–
11
14
21
–
28
11
25
23
11
17
17
–
29
15
22
27
11
12
13
Ciprofloxacin
Gatifloxacin
Table 2. Minimal inhibitor concentration (MIC) mg/mL of the synthesized compounds 2–13 against the tested bacterial strains
Comp. no.
Pseudomanas
aeruginosa
ATCC 27853
Klebsiella
pneumoniae
CIP 53153
Escherichia
coli
ATCC 25922
Staphylococcus
aureus
ATCC 25923
2
3
4
5
6
7
8
9
10
–
60
150
180
–
300
–
180
130
–
–
150
30
150
240
180
60
120
150
150
15
–
–
270
120
60
90
120
30
15
–
0.9375
120
Nil
–
120
60
15
–
3.75
–
Nil
15
7.5
120
–
15
11
12
13
120
3.75
60
Nil
7.5
1.875
120
Nil
3.75
3.75
Control
Ciprofloxacin
Gatifloxacin
3.75
1.875
3.75
bacterial activity against Gram-positive bacteria S. aureus
ATCC 25923 with lowest MIC of 0.9375 mg/mL as compared
to the standard drugs ciprofloxacin (MIC 3.75 mg/mL) and
gatifloxacin (MIC 1.875 mg/mL). The newly synthesized com-
pounds were also tested for approximate lethal dose LD50 and
were found to exhibit a higher value of LD50, that is, more than
800 mg/kg, except compound 12 which exhibited LD50 of more
than 1600 mg/kg (maximum dose tested). The compounds
have shown high value of LD50 indicating good safety margin.
Initial anticonvulsant activity and neurotoxicity (NT) data
for the compounds 2–13 are reported in Table 3, along with
the literature data on phenytoin, carbamazapine, sodium
valproate, phenobarbital, and ethosuximide. All the phos-
phazene compounds showed more effective sedative-hyp-
notic and central nervous system (CNS)-depressant activity
rather than anticonvulsant activity. In the prepared series 4,
5, and 11 were found to be active in the maximal electroshock
seizure (MES) screen at 0.5 h, whereas 11 showed anticonvul-
sant activity at 4 h. All the compounds exhibited activity at
300 mg/kg body weight. Except 4, 5, and 11, all the remaining
compounds were inactive in the scPTZ screen at 0.5 h. Only
11 showed activity at both 0.5 and 4 h interval time which
indicates their rapid action and long duration of activity.
Synthesized compounds exhibited NT at only 300 mg/kg. 4
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E. Cil et al.
Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
Table 3. Anticonvulsant activity and minimal motor impairment of the compounds 2–13
Comp. no.
Intraperitoneal injection in micea)
ScPTZ screen
MES screen
Neurotoxicity screen
0.5 h
4 h
0.5 h
4 h
0.5 h
4 h
2
3
–
–
–
–
–
–
–
–
300
–
–
–
4
5
6
300
300
–
–
–
–
300
300
–
–
–
–
300
300
–
300
–
–
7
–
–
–
–
–
–
8
–
–
–
–
–
–
9
10
–
–
–
–
–
–
–
–
–
–
–
–
11
12
300
–
300
–
300
–
–
–
–
–
–
–
13
–
–
–
–
100
300
30
300
–
–
300
–
300
–
300
100
100
–
100
–
300
100
300
–
300
–
Phenytoin
Carbamazepine
Sodium valproate
Phenobarbitals
Ethosuximide
30
30
–
100
–
30
100
–
30
–
a)
Doses of 30, 100, and 300 mg/kg were administered. The figures in the table indicate the minimum dose whereby bioactivity was
demonstrated in half or more of the mice. The animals were examined 0.5 and 4 h after injections. – indicates an absence of activity at
maximum dose administered (300 mg/kg).
and 13 showed extended NT lasting up to 4 h. The NT of the
compounds as indicated by their inability to grasp the rotat-
ing rot for sufficient time may be due to the increase blood–
brain barrier (BBB) penetration and accumulation at neuro-
protein sites and then binding to unwanted receptor sites. All
the compounds were also screened for behavior study and
CNS-depressant activity. In the behavioral study using acto-
photometer scoring technique, the entire synthesized com-
pounds showed decrease in locomotor activity where 42%
was the lowest and 53% was the maximal decrease in loco-
motor activity when compared to phenytoin as reported in
Table 4. All the compounds except 12 showed more than 44%
Table 4. Behavioral study of the compounds 2–13
Compounda)
Activity scoreb)
0.5 h after
% Inhibition
Control (24 h prior)
Post-treatment
1 h after
2
3
4
5
6
7
8
9
10
483.19 ꢁ 0.72
484.21 ꢁ 0.34
323.12 ꢁ 0.84
514.49 ꢁ 0.83
475.26 ꢁ 1.28
415.48 ꢁ 0.66
530.34 ꢁ 0.85
401.21 ꢁ 0.98
521.32 ꢁ 1.19
481.02 ꢁ 1.24
490.17 ꢁ 0.79
495.72 ꢁ 0.31
645.98 ꢁ 31.11
289.10 ꢁ 0.78
322.65 ꢁ 1.32
311.98 ꢁ 0.83
297.70 ꢁ 0.92
274.08 ꢁ 0.99
390.98 ꢁ 1.32
298.05 ꢁ 0.47
281.95 ꢁ 0.78
327.30 ꢁ 2.24
274.03 ꢁ 0.99
343.11 ꢁ 1.18
321.15 ꢁ 1.32
251.13 ꢁ 12.45
242.20 ꢁ 0.19
261.82 ꢁ 0.85
121.04 ꢁ 0.87
251.72 ꢁ 0.21
221.12 ꢁ 1.28
209.02 ꢁ 1.40
266.91 ꢁ 0.22
187.82 ꢁ 0.99
249.41 ꢁ 0.30
223.45 ꢁ 1.22
284.49 ꢁ 1.26
273.15 ꢁ 0.86
194.21 ꢁ 30.15
50
45
48
51
53
40
51
51
52
53
42
45
70
11
12
13
Phenytoinc)
a)
The compounds were tested at a dose of 100 mg/kg (i.p.).
Each score represents the mean ꢁ SEM of six mice, significantly different from the control score at <0.02 and NS indicates not
b)
significant at p >0.02 (Student’s t-test).
c)
Tested at 25 mg/kg i.p.
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Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
Synthesis, Characterization, and Biological–Pharmacological Evaluation of New Phosphazenes
7
Table 5. CNS study on the compounds 2–13
tive sedative-hypnotic and CNS-depressant activity as com-
pared to anticonvulsant activity. Bulkier compounds are
more lipophilic and can cross the BBB to exert their effect
at the CNS. Substituted phosphazene compounds (2–13) have
larger molecular structure. So, it may also be concluded that
because of this larger molecular structure, substituted phos-
phazene compounds (2–13) possess good CNS activity.
Compounda)
Immobility timeb) (s)
Control
(24 h prior)
Post-treatment
(60 min after)
PEG
2
3
4
5
6
7
8
9
10
11
12
13
173.16 ꢁ 10.70
73.21 ꢁ 18.48
121.00 ꢁ 17.10
130.72 ꢁ 18.34
45.60 ꢁ 19.42
130.12 ꢁ 15.12
176.78 ꢁ 17.82
181.12 ꢁ 14.35
173.07 ꢁ 17.86
174.75 ꢁ 13.28
161.98 ꢁ 17.58
177.92 ꢁ 18.52
151.10 ꢁ 8.43
178.15 ꢁ 14.32 NS
130.65 ꢁ 11.52
188.61 ꢁ 13.23
204.92 ꢁ 12.95
70.36 ꢁ 14.72
Experimental
193.98 ꢁ 14.61
190.92 ꢁ 12.88 NS
225.60 ꢁ 16.68
218.32 ꢁ 21.27
211.32 ꢁ 20.32
209.30 ꢁ 21.12
203.30 ꢁ 13.15 NS
198.32 ꢁ 16.12
240.35 ꢁ 14.18
General remarks
Solvents and other liquids used in the experimental works were
dried by conventional methods. Hexachlorocyclotriphosphazene
[N3P3Cl6] (1) was recrystallized from hexane. Other chemicals
were used as purchased. 2,2-Dichloro-4,4,6,6-bis[spiro(20,200-
dioxy-10,100-biphenylyl]cyclotriphoshazene (2) was prepared as
described by Carriedo et al. [30]. The reaction of [N3P3Cl6] with
the biphenyl-2,20-diol was carried out under dry nitrogen. IR
spectra were recorded on an ATI Unicam Mattson 1000 FTIR
spectrometer. 1H, 13C, and 31P NMR spectra were recorded using
a Bruker DPX-300 spectrometer operating at 300.13, 75.46, and
121.49 MHz, respectively. The 1H and 13C NMR chemical shifts
were measured using SiMe4 as an internal standard, whereas
those for 31P were measured using 85% H3PO4 as an external
standard. For the NMR studies, the CDCl3-d was used as solvent
(CDCl3-d þ DMSO-d were used for 9). Chemical shifts downfield
from the standard were assigned positive d values. Microanalysis
was carried out by a LECO 932 CHNS-O apparatus.
Carbamazepinec) 138.11 ꢁ 15.28
a)
The compounds were tested at a dose of 100 mg/kg (i.p.).
Each value represents the mean ꢁ SEM of six rats significantly
b)
different from the control at p <0.05 and NS denotes not sig-
nificant at p <0.05 (Student’s t-test).
c)
Tested at 30 mg/kg (i.p).
decrease in locomotor activity (p <0.05) after 1 h of com-
pound administration. In a similar study with forced swim
pool test, the immobility time after administration of the test
compounds was compared with carbamazepine (Table 5).
Readings of control groups were taken individually for each
compound 24 h prior to compound administration.
Experimental results indicate that our compounds exhibited
better sedative-hypnotic and CNS-depressant activity as com-
pared to anticonvulsant activity. Biological activity was also
ascertained for PEG because it was used as a vehicle for the
synthesized compounds. Except for 7 and 12 other tested
compounds were found to exhibit potent CNS-depressant
activity as indicated by increased immobility time.
Chemistry
Synthesis of compound 2
A mixture of 1 (10.20 g, 29.34 mmol), biphenyl-2,20-diol (10.70 g,
57.46 mmol), and K2CO3 (20.00 g, 144.70 mmol) was stirred in
acetone (100 mL) at 08C and then was reacted at ambient
temperature for 24 h. The solvent was removed under vacuum.
The residue was extracted with CH2Cl2 (4 ꢂ 75 mL). After the
solvent was removed, a white solid (2) formed and crystallized
from acetone (15.48 g, 92%). Anal. Calc. for C24H16Cl2N3O4P3
(MW ¼ 574.22): C, 50.20; H, 2.81; N, 7.32; found: C, 49.80; H,
2.70; N, 7.00%. IR (KBr/cmꢀ1): 3034 and 3071 nC–H(ar.), 1194
nP–N, 942 nP–O–C.
1H NMR: 7.68 (4 H, d, J 7.45, H5); 7.55 (4 H, t,
–
J 7.60, H3); 7.40 (8 H, m, H2, H4). 13C NMR: 147.30 (2JPOC 8.86, C1);
130.73 (C5); 130.52 (C3); 129.00 (C6); 126.99 (C4); 121.96 (C2). 31P
NMR: AX2 system, 26.60 (t, A port.), 19.90 (d, B port.).
Conclusion
For the preparation of the phosphazenes containing Schiff
base (with high yield), this method is very suitable. The
compounds 2–13 exhibited varying antibacterial response
against different types of bacterial strains. However, the
compounds having groups such as OH, Cl, and CN showed
better results than the other compounds. Especially, com-
pound 12 having both Cl and OH groups showed promising
antibacterial activity against both types of bacteria. All the
compounds exhibited LD50 >800 mg/kg, except compound
12 (having Cl and OH groups) which showed LD50 >1600 mg/
kg. Based upon the experimental results it may be concluded
that the newly synthesized compounds showed more effec-
Synthesis of compound 3
A mixture of 2 (15 g, 26.12 mmol), 2-hydroxybenzaldehyde
(6.70 g, 54.80 mmol), and K2CO3 (21.00 g, 151.94 mmol) was
stirred in THF (100 mL) at 08C and then refluxed for 12 h. The
solvent was removed under vacuum. The residue was extracted
with CH2Cl2 (4 ꢂ 75 mL). The solvent was removed and the
product was dissolved in a small amount of chloroform and was
precipitated with hexane several times. The white solid (3) was
washed with ether and dried at 508C under vacuum. Yield: 81%
(15.77 g). Anal. Calc. for C38H26N3O8P3 (MW ¼ 745.55): C, 61.22;
H, 3.52; N, 5.64. Found: C, 61.38; H, 3.28; N, 5.58%. IR (cmꢀ1): 1694
nC–O, 1171 nP–N, 938 nP–O–C. 1H NMR: 10.44 (2H, s, H13); 8.06 (2H, d,
–
–
J
8.32,
H
11); 7.05 (2H, t, 7.52, H10); 7.33–7.73 (20
J
H, m, H2, H3, H4, H5, H8, H9). 13C NMR: 188.69 (C13); 152.60
ß 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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8
E. Cil et al.
Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
(C7); 147.85 (C1); 135.44 (C9); 129.80 (C5); 129.69 (C3); 128.62 (C11);
128.55 (C10); 128.48 (C6); 126.26 (C4); 126.13 (C12); 122.66 (C8);
121.69 (C2). 31P NMR: AX2 system, 9.39 (t, A port.), 24.99 (d, B port.).
6.95, H14); 1.60 (6H, d, J 7.60, H25); 6.80–7.29 (26 H, m, H2, H3,
H4, H5, H8, H9, H10, H11, H16, H17, H19, H20, H21, H22, H24). 13C NMR:
166.73 (C13); 154.40 (C7); 139.16 (C1); 133.15 (C9); 132.89–124.65
(19C, C2, C3, C4, C5 C6, C8, C10, C11, C12, C15, C16, C17, C18, C19,
C20, C21, C22, C23, C24); 51.20 (C14); 22.25 (C25). 31P NMR: AX2
system, 10.23 (t, A port.), 25.09 (d, B port.).
General method for synthesis of compounds 4–13
Schiff base compounds were prepared according to the known
method from the condensation of compound 3 with amines in
mole ratio of 1:2. The mixture in THF was reacted by refluxing for
48 h. After the reaction was completed, the solvent was evapor-
ated under reduced pressure. The crude product was dissolved in
a small amount of chloroform and was precipitated with hexane
several times. And finally it was washed with ether and dried at
508C under vacuum.
Synthesis of 8
Compound 3 (0.60 g, 0.80 mmol) and 2-aminophenol (0.17 g,
1.62 mmol) were used. Yield: 87% (0.65 g). Yellow solid. Anal.
Calc. for C50H36N5O6P3 (MW ¼ 927.77): C, 64.73; H, 3.91; N, 7.55.
Found: C, 64.56; H, 3.89; N, 7.52%. IR (cmꢀ1): 3422 nO–H, 1621 nC–
,
N
–
1168 nP–N, 944 nP–O–C. 1H NMR: 9.09 (2H, s, H20); 8.33 (2H, s, H13); 6.64
–
(2H, d, J 7.80, H18); 6.95–7.64 (30 H, m, H2, H3, H4, H5, H8, H9,
H
10, H11, H15, H16 H17). 13C NMR: 152.37 (C13); 152.29 (C7); 150.69
Synthesis of 4
(C19); 147.83 (C1); 135.61 (C9); 132.54 (C14); 129.73 (C5); 129.61 (C3);
128.96 (C11); 128.61 (C10); 128.55 (C17); 127.88 (C6); 126.15 (C4);
126.10 (C12); 122.63 (C8); 121.68 (C2); 120.10 (C15); 116.48 (C18);
114.80 (C16). 31P NMR: AX2 system, 10.60 (t, A port.), 25.06 (d, B
port.).
Compound 3 (0.60 g, 0.80 mmol) and 4-methoxybenzylamine
(0.22 g, 0.21 mL, 1.60 mmol) were used. Yield: 81% (0.64 g).
Fawn colored solid. Anal. Calc. for C54H44N5O8P3 (MW ¼ 938.87):
C, 65.92; H, 4.51; N, 7.12. Found: C, 65.83; H, 4.37; N, 7.18%. IR
(cmꢀ1): 1657 nC–N, 1176 nP–N, 940 nP–O–C. 1H NMR: 8.60 (2H, s, H13);
–
–
7.53–6.74 (32 H, m, H2, H3, H4, H5, H8, H9, H10, H11, H16, H17); 4.37
(4H, s, H14); 3.81 (6H, s, H19). 13C NMR: 168.82 (C18); 161.05 (C13);
156.86 (C7); 147.94 (C1); 131.72 (C9); 131.28 (C15); 130.34 (C16); 129.80
(C5); 129.58 (C3); 129.16 (C11); 128.71 (C10); 128.62 (C6); 126.15 (C4);
125.96 (C12); 125.80 (C8); 121.85 (C2); 114.10 (C17); 66.63 (C14); 55.26
(C19). 31P NMR: AX2 system, 10.46 (t, A port.), 25.28 (d, B port.).
Synthesis of 9
Compound 3 (0.60 g, 0.80 mmol) and 4-aminophenol (0.17 g,
1.62 mmol) were used. Yield: 90% (0.67 g). Yellow solid. Anal.
Calc. for C50H36N5O6P3 (MW ¼ 927.77): C, 64.73; H, 3.91; N, 7.55.
Found: C, 64.70; H, 3.89; N, 7.50%. IR (cmꢀ1): 3341 nO–H, 1619 nC–
,
N
–
1166 nP–N, 945 nP–O–C. 1H NMR: 8.77 (2H, s, H18); 8.27 (2H, s, H13);
–
6.51 (2H, d, J 7.83, H16); 6.62–7.74 (30 H, m, H2, H3, H4, H5, H8,
H9, H10, H11, H15). 13C NMR: 156.66 (C13); 151.52 (C7); 147.58 (C1);
143.17 (C17); 131.89 (C9); 130.00 (C5); 129.68 (C3); 129.12 (C11);
128.31 (C10); 127.64 (C6); 126.44 (C4); 126.16 (C12); 122.58 (C8);
122.16 (C14); 121.75 (C2); 116.22 (C15); 115.94 (C16). 31P NMR: AX2
system, 10.83 (t, A port.), 24.97 (d, B port.).
Synthesis of 5
Compound 3 (0.60 g, 0.80 mmol) and 2-chlorobenzylamine
(0.23 g, 0,20 mL, 1.62 mmol) were used. Yield: 73% (0.58 g).
Yellow solid. Anal. Calc. for C52H38Cl2N5O6P3 (MW ¼ 992.71): C,
62.91; H, 3.86; N, 7.05. Found: C, 63.00; H, 3.81; N, 7.13%. IR
(cmꢀ1): 1659 nC–N, 1174 nP–N, 944 nP–O–C. 1H NMR: 8.36 (2H, s, H13);
–
–
5.41 (4H, s,
H
14); 6.95–7.66 (32 H, m, H2, H3, H4,
H5, H8, H9, H10, H11, H16, H17 H18, H19). 13C NMR: 166.04 (C13);
161.60 (C7); 147.65 (C1); 136.91 (C9); 133.39 (C15); 132.11 (C20);
130.10 (C18); 129.99 (C5); 129.76 (C3); 129.47 (C19); 129.44 (C17);
129.33 (C16); 128.69 (C11); 128.43 (C10); 128.21 (C6); 126.90 (C4);
126.50 (C12); 126.13 (C8); 121.73 (C2); 61.94 (C14). 31P NMR: AX2
system, 10.13 (t, A port.), 25.09 (d, B port.).
Synthesis of 10
Compound 3 (0.60 g, 0.80 mmol) and 4-aminobenzonitrile
(0.19 g, 1.62 mmol) were used. Yield: 79% (0.51 g). Yellow solid.
Anal. Calc. for C45H30N5O7P3 (MW ¼ 845.67): C, 63.91; H, 3.58; N,
8.28. Found: C, 63.84; H, 3.49; N, 8.25%. IR (cmꢀ1): 1692 nC–O, 1620
–
nC–N, 1171 nP–N, 940 nP–O–C
–
(1H, s,
.
1H NMR: 10.44 (1H, s, H24); 8.62
–
H J
13); 8.30 (1H, d, 7.92, H22); 6.90–7.52 (27
H, m, H2, H3, H4, H5, H8, H9, H10, H11, H15, H16, H19, H20, H21).
13C NMR: 188.69 (C24); 157.86 (C13); 155.98 (C14); 151.15 (C7);
150.71 (C17); 150.41 (C18); 147.81 (C1); 147.66 (C16); 135.40 (C9);
133.35 (C21); 129.79 (C5); 129.64 (C3); 128.49 (C11); 128.46 (C22);
128.36 (C10); 128.31 (C20); 128.11 (C6); 126.31 (C4); 126.18 (C23);
126.12 (C12); 122.89 (C8); 122.79 (C19); 121.64 (C2); 120.12 (C25);
119.03 (C15). 31P NMR: AX2 system, 11.10 (t, A port.), 24.98 (d, B
port.).
Synthesis of 6
Compound 3 (0.60 g, 0.80 mmol) and 4-chlorobenzylamine
(0.23 g, 0,20 mL, 1.62 mmol) were used. Yield: 80% (0.64 g).
Yellow solid. Anal. Calc. for C52H38Cl2N5O6P3 (MW ¼ 992.71): C,
62.91; H, 3.86; N, 7.07. Found: C, 62.79; H, 3.95; N, 6,92%. IR (cmꢀ1):
1659 nC–N, 1174 nP–N, 943 nP–O–C.
1H NMR: 8.17 (2H, s, H13); 4.05
–
–
(4H, s, H14); 6.94–7.67 (32 H, m, H2, H3, H4, H5, H8, H9,
H
10, H11, H16, H17). 13C NMR: 166.09 (C13); 161.65 (C7); 147.33
(C1); 133.49 (C9); 130.22 (C18); 129.91 (C5); 129.71 (C3); 129.58
(C17); 129.50 (C16); 129.28 (C15); 129.03 (C11); 128.68 (C10); 128.44
(C6); 127.37 (C4); 127.05 (C12); 126.44 (C8); 121.69 (C2); 42.30 (C14).
31P NMR: AX2 system, 10.13 (t, A port.), 25.09 (d, B port.).
Synthesis of 11
Compound 3 (0.60 g, 0.80 mmol) and biphenyl-2-amine (0.23 g,
1.62 mmol) were used. Yield: 82% (0.43 g). Yellow solid. Anal.
Calc. for C62H44N5O6P3 (MW ¼ 1047.96): C, 71.06; H, 4.23; N, 6.68.
Found: C, 71.12; H, 4.40; N, 6.67%. IR (cmꢀ1): 1622 nC–N, 1171 nP–
,
–
N
Synthesis of 7
–
.
1H NMR: 8.76 (2H, s, H13); 8.29 (2H, d, J 8.15, H19);
Compound 3 (0.60 g, 0.80 mmol) and 2-naphthylethylamine
(0.27 g, 1.62 mmol) were used. Yield: 84% (0.71 g). Orange solid.
Anal. Calc. for C52H38Cl2N5O6P3 (MW ¼ 1052.0): C, 70.79; H, 4.60;
943 nP–O–C
6.70–7.71 (40 H, m, H2, H3, H4, H5, H8, H9, H10, H11, H16, H17, H18
,
H
21, H22, H23). 13C NMR: 154.95 (C13); 150.35 (C14); 150.28 (C7);
N, 6.66. Found: C, 70.65; H, 4.56; N, 6.66%. IR (cmꢀ1): 1654 nC–
,
N
147.62 (C1); 139.42 (C20); 135.42 (C9); 132.37 (C18); 130.11 (C15);
130.02 (C16); 129.92 (C23); 129.79 (C5); 129.66 (C3); 129.08 (C22);
–
1173 nP–N, 949 nP–O–C.
1H NMR: 8.44 (2H, s, H13); 4.14 (4H, q, J
–
ß 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
Synthesis, Characterization, and Biological–Pharmacological Evaluation of New Phosphazenes
9
129.03 (C21); 128.52 (C11); 128.31 (C10); 128.12 (C6); 127.64 (C19);
126.80 (C17); 126.41 (C4); 126.24 (C12); 122.32 (C8); 121.64 (C2).
31P NMR: AX2 system, 10.83 (t, A port.), 25.14 (d, B port.).
lowest concentrations of compound that completely inhibited
the growth of each strain. All compounds dissolved in dimethyl-
sulfoxide were added to the culture media. Mueller–Hinton
Broth for bacteria was used to obtain the final concentrations
ranging from 300 to 0.9375 mg/mL. The amounts of dimethylsulf-
oxide never exceed 1% v/v. Inocula consisted of 5.0 ꢂ 104 bacteria/
mL. The MICs were read after incubation at 378C for 24 h (bacteria).
Media and media with 1% v/v dimethylsulfoxide were employed
as growth controls. Ciprofloxacin and gatifloxacin were used
as reference antibacterial activity, subcutaneous were performed
by transferring 100 mL of each mixture remaining clear in 1 mL
of fresh medium. The minimal bactericidal concentrations (MBC,
mg/mL) were read after incubation at 378C for 48 h.
Synthesis of 12
Compound 3 (0.60 g, 0.80 mmol) and 2-amino-4-chlorophenol
(0.23 g, 1.62 mmol) were used. Yield: 85% (0.45 g). Yellow solid.
Anal. Calc. for C50H34Cl2N5O8P3 (MW ¼ 996.60): C, 60.25; H, 3.44;
N, 7.03. Found: C, 60.50; H, 3.38; N, 7.28%. IR (cmꢀ1): 3370 nO–H
,
1620 nC–N, 1168 nP–N, 945 nP–O–C. 1H NMR: 10.50 (2H, s, H20); 8.85
–
–
(2H, s, H13); 8.25 (2H, d, J 7.56, H11); 6.42 (2H, d, J 8.10, H16); 6.82–
7.70 (36 H, m, H2, H3, H4, H5, H8, H9, H10, H11, H17, H19). 13C NMR:
153.70 (C13); 150.93 (C15); 150.66 (C7); 150.58 (C14); 147.75 (C1);
136.42 (C9); 133.00 (C18); 129.70 (C5); 129.66 (C3); 128.53 (C11);
128.50 (C10); 128.44 (C6); 128.16 (C17); 126.24 (C4); 126.17 (C12);
122.60 (C8); 121.60 (C2); 116.62 (C19); 115.88 (C16). 31P NMR: AX2
system, 10.78 (t, A port.), 24.83 (d, B port.).
Approximate lethal dose (LD50
)
The compounds were investigated for this acute toxicity (LD50) in
albino mice by following the method of Smith [45]. Test com-
pounds were administered orally in one group and the same
volume of normal saline in another group of animals consisting
of six mice each in graded doses. During the study, the animals
were allowed to take water and food ad libitum. After 24 h of drug
administration percent mortality in each group was observed.
From the data obtained LD50 was calculated.
Synthesis of 13
Compound 3 (0.60 g, 0.80 mmol) and pyrimidine-2-amine
(0.15 g, 1.62 mmol) were used. Yield: 78% (0.53 g). Yellow solid.
Anal. Calc. for C42H29N6O7P3 (MW ¼ 822.64): C, 61.32; H, 3.55; N,
10.22. Found: C, 61.68; H, 3.80; N, 10.35%. IR (cmꢀ1): 1698 nC–
,
O
–
1619 nC–N, 1171 nP–N, 945 nP–O–C. 1H NMR: 10.43 (1H, s, H23); 8.31
–
–
(2H, d, J 8.15, H15); 8.06 (1H, d, J 7.90, H13); 7.04–7.70 (25
H, m, H2, H3, H4, H5, H8, H9, H10, H11, H16, H18, H19, H20, H21).
13C NMR: 188.72 (C23); 161.20 (C13); 158.30 (C15); 157.86 (C14);
152.61 (C7); 152.54 (C17); 147.86 (C1); 135.50 (C9 and C20); 129.83
(C5); 129.61 (C3); 128.62 (C11); 128.53 (C21); 128.47 (C10 and C19);
128.19 (C6); 126.28 (C4); 128.17 (C12); 125.81 (C22); 122.66 (C8);
121.84 (C18); 121.68 (C2); 111.51 (C16). 31P NMR: AX2 system, 10.78
(t, A port.), 24.85 (d, B port.).
Pharmacology
The new derivatives obtained from the reaction sequence were
injected intraperitoneally into mice and evaluated in the MES,
scPTZ, and NT screens, using doses of 30, 100, and 300 mg/kg
at two different time intervals. These data are presented in
Table 3. These compounds were also screened for their
CNS behavioral activity in mice using actophotometer, CNS-
depressant activity with the help of the forced swim method.
The data of the pharmacological studies are reported in
Tables 3–5.
Biological studies
Antibacterial activity
Anticonvulsant screening
The newly synthesized compounds 2–13 were screened for anti-
bacterial activity against bacterial strains namely P. aeruginosa
ATCC 27853, K. pneumoniae CIP 53153, E. coli ATCC 25922, and S.
aureus ATCC 25923 at a concentration of 300 mg/mL by the filter
paper disc method [43]. For comparison, ciprofloxacin and gati-
floxacin were used as the standard drugs. DMSO served as control
and there was no visible change in bacterial growth due to this.
The discs of Whatman filter paper were prepared with standard
size (7 mm) and kept in one-Oz screw-capped wide-mouthed
containers for sterilization. These bottles are kept in the hot-
air oven at 1508C. Now, solution is put into each bottle. The discs
are transferred to the inoculated plates with a pair of fine
pointed tweezers. To prevent contamination, tweezers may be
kept with their tips in 70% alcohol and flamed off before use.
Before using the test organisms, grown on nutrient agar, they
were subcultured on nutrient broth at 378C for 18–20 h. Each
disc was carefully applied to the surface of the agar without
lateral movement once the surface had been touched. Now, the
plates were incubated for 24 h at 378C.
Initially all the compounds were administered i.p. in a volume of
0.01 mL/g body weight for mice and 0.004 mL/g body weight for
rats at doses of 30, 100, 300 mg/kg to one to four animals.
Activity was established using the MES and scPTZ test and these
data are presented in Table 3.
Neurotoxicity screen
Minimal motor impairment was measured in mice by the
rotorod test. The mice were trained to stay on an accelerating
rotorod that rotated at 6 rpm. The rod diameter was 3.2 cm.
NT was indicated by the inability of the animal to maintain
equilibrium on the rod for at least 1 min in each of the three
trials.
Behavioral testing
The title compounds (100 mg/kg) were screened for their behav-
ioral effect using actophotometer [46] at 30 min and 1 h after
drug administration. The behavior of animals inside the photo-
cell was recorded as a digital score. Increased scores suggest good
behavioral activity. The activity of the compounds was at maxi-
mum at 1 h; therefore, the activity values at 1 h were used to
calculate % decrease in locomotor activity. The control group
animals were administered PEG 400. The observations are tabu-
lated in Table 4.
Minimal inhibitory concentration (MIC)
The antibacterial activity was assayed in vitro by twofold broth
dilution [44] against the bacterial strains P. aeruginosa ATCC
27853, K. pneumoniae CIP 53153, E. coli ATCC 25922, and
S. aureus ATCC 25923. The MIC (in mg/mL) were defined as the
ß 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com

10
E. Cil et al.
Arch. Pharm. Chem. Life Sci. 2012, 000, 1–10
[19] E. Cil, M. Arslan, F. Aslan, A. I. Ozturk, Phosphorus, Sulfur
Silicon Relat. Elem. 2003, 178, 1037–1046.
CNS-depressant activity
The forced swim pool method described earlier [47] was followed.
Wistar rats were placed in a chamber (diameter 45 cm and height
20 cm) containing water up to a height of 15 cm at 25 ꢁ 28C.
Two swim sessions were conducted an initial 15 min pre-test,
followed by a 5-min test session 24 h later. The animals were
drug administrated (100 mg/kg) the test compound i.p. 30 min
before the test session. The period of immobility (passive floating
without struggling, making only those movements which are
necessary to keep its head above the surface of water) during the
5 min test period was measured. The results are presented in
Table 5.
[20] A. I. Ozturk, F. Aslan, E. Cil, M. Arslan, Phosphorus, Sulfur
Silicon Relat. Elem. 2003, 178, 2097–2105.
[21] M. Arslan, F. Aslan, A. I. Ozturk, Heteroat. Chem. 2003, 14, 138–143.
[22] F. Aslan, A. I. Ozturk, M. Arslan, Heteroat. Chem. 2003, 14, 413–
416.
[23] T. Rodima, I. Kaljurand, A. Pihl, V. Maemets, I. Leito, I. A.
Koppel, J. Org. Chem. 2002, 67, 1873–1881.
[24] R. Bertani, G. Facchin, M. Gleria, Inorg. Chim. Acta 1989, 165,
73–82.
[25] A. Kılıc¸, S. Begec¸, Z. Kılıc¸, T. Hokelek, J. Mol. Struct. 2000, 516,
255–262.
We thank the Firat University Research Fund for support.
The authors have declared no conflict of interest.
[26] G. A. Carriedo, L. F. Catuxo, F. J. G. Alonso, P. G. Elipe, P. A.
Gonzalez, G. Sanchez, J. Appl. Polym. Sci. 1996, 59, 1879–1885.
[27] Z. Ngaini, A. R. I. Norashikin, Phosphorus, Sulfur Silicon Relat.
Elem. 2010, 185, 628–633.
References
[28] S. Begec, A. Kılıc¸, Heteroat. Chem. 2005, 16, 308–310.
[29] D. Kumar, A. D. Gupta, Macromolecules 1995, 28, 6323–6329.
[1] H. R. Allcock, Phosphorus–Nitrogen Compounds, Academic
Press, New York 1972, pp. 1–52.
[30] G. A. Carriedo, L. F. Catuxo, F. J. G. Alonso, P. G. Elipe, P. A.
Gonzalez, Macromolecules 1996, 29, 5320–5325.
[2] G. A. Carriedo, F. J. G. Alonso, J. L. Garcia, R. J. Carbajo, F. L.
Ortiz, Eur. J. Inorg. Chem. 1999, 6, 1015–1020.
[31] U. Belluco, R. Bertani, R. A. Michelin, M. Mozzan, F. Zingales,
M. Gleria, Inorg. Chim. Acta 1995, 229, 13–15.
[3] P. J. Lieu, J. H. Magill, Y. C. Alarie, J. Fire Flammability 1980, 11,
167–183.
[32] M. B. McIntosh, T. J. Hartle, H. R. Allcock, J. Am. Chem. Soc.
1999, 121, 884–885.
[4] M. A. Olshavsky, H. R. Allcock, Macromolecules 1995, 28, 6188–
6197.
[33] F. Palacios, C. Alonso, G. G. Rubialesa, J. M. Ezpeleta,
Tetrahedron 2004, 60, 2469–2474.
[5] H. R. Allcock, A. A. Dembek, C. Kim, R. L. S. Devine, Y. Shi,
W. H. Steier, C. W. Spangler, Macromolecules 1991, 24, 1000–
1010.
[34] E. Cil, M. Arslan, A. O. Gorgulu, Heteroat. Chem. 2006, 17, 112–
117.
[35] E. Cil, M. Arslan, A. O. Gorgulu, Can. J. Chem. 2005, 83, 2039–
2045.
[6] A. Dembek, C. Kim, H. R. Allcock, R. L. S. Devine, Y. Shi, W. H.
Steier, C. W. Spangler, Chem. Mater. 1991, 2, 97–99.
[36] O. Pamukci, E. Cil, S. Begec¸, M. Arslan, Heteroat. Chem. 2007,
18, 791–797.
[7] H. R. Allcock, C. Kim, Macromolecules 1991, 24, 2846–2851.
[8] F. Minto, M. Gleria, P. Bortolus, L. Fambri, A. Pegoretti,
J. Appl. Polym. Sci. 1995, 56, 747–756.
[37] E. Cil, M. Arslan, Inorg. Chim. Acta 2009, 362, 1421–1427.
[38] W. Yuan, L. Zhu, X. Huang, S. Zheng, X. Tang, Eur. Polym. J.
2005, 41, 1867–1873.
[9] C. T. Laurencin, H. J. Koh, T. X. Neenan, H. R. Allcock,
R. Langer, J. Biomed. Mater. Res. 1987, 21, 1231–1246.
[39] W. Yuan, X. Tang, X. Huang, S. Zheng, Polymer 2005, 46,
1701–1707.
[10] H. R. Allcock, S. R. Pucher, R. J. Fitzpatrick, K. Rashid,
Biomaterials 1992, 13, 857–862.
[40] G. Fontana, F. Minto, M. Gleria, G. Facchin, R. Bertani, G.
Favero, Eur. Polym. J. 1996, 32, 1273–1283.
[11] G. Giavaresi, M. Tschon, V. Borsari, J. H. Daly, J. J. Liggat,
M. Fini, V. Bonazzi, A. Nicolini, A. Carpi, M. Morra, C.
Cassinelli, R. Giardino, Biomed. Pharmacother. 2004, 58,
411–417.
[41] T. Muraki, M. Ueta, E. Ihara, K. Inoue, Polym. Degrad. Stab.
2004, 84, 87–93.
[42] F. Aslan, Z. Demirpence, R. Tatsiz, H. Turkmen, A. I. Ozturk,
M. Arslan, Z Anorg. Allg. Chem. 2008, 634, 1140–1144. and
references cited therein.
[12] Y. J. Jun, J. I. Kim, M. J. Jun, Y. S. Sohn, J. Inorg. Biochem. 2005,
99, 1593–1601.
[13] K. Brandt, R. Kruszynski, T. J. Bartczak, I. P. Czomperlik,
Inorg. Chim. Acta 2001, 322, 138–144.
[43] J. C. Gould, J. H. Bowie, Edinb. Med. J. 1952, 59, 178–179.
[44] J. H. Jorgersen, J. D. Turnidge, J. A. Washington, in Manual of
Clinical Microbiology. (Eds: P. R. Murray, E. J. Baron, M. A.
Pfaller, F. C. Tenover, R. H. Nolken), American Society for
Microbiology, Washington, DC 1999, pp. 1275–1434.
[14] A. I. Ozturk, O. Yılmaz, S. Kırbag, M. Arslan, Cell Biochem.
Funct. 2000, 18, 117–126.
[15] V. Konar, O. Yılmaz, A. I. Ozturk, S. Kırbag, M. Arslan, Bioorg.
Chem. 2000, 28, 214–225.
[45] Q. E. Smith, Pharmacological Screening Tests Progress in Medicinal
Chemistry, vol. 1, Butterworth, London 1960, pp. 1–33.
[16] O. Yılmaz, F. Aslan, A. I. Ozturk, N. S. Vanlı, S. Kırbag,
M. Arslan, Bioorg. Chem. 2002, 30, 303–314.
[46] J. R. Boissoer, P. Simon, Arch. Int. Pharmacodyn. Ther. 1965, 158,
212–214.
[17] M. Gleria, R. D. Jaeger, J. Inorg. Organomet. Polym. 2001, 11,
1–45.
[47] R. D. Porsolt, G. Anton, N. Blanet, M. Jalbre, Eur. J. Pharmacol.
1978, 47, 379–391.
[18] M. T. R. Laguna, J. Gallego, F. Menduciti, E. Saiz, M. P.
Tarazona, Macromolecules 2002, 35, 7782–7790.
ß 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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