Synthesis, anti-inflammatory, analgesic and kinase (CDK-1, CDK-5 and GSK-3) inhibition activity evaluation of benzimidazole/benzoxazole derivatives and some Schiff's bases

Article abstract of DOI:10.1016/j.bmc.2006.01.054

A series of N-(acridin-9-yl)-4-(benzo[d]imidazol/oxazol-2-yl) benzamides has been synthesized by the condensation of 9-aminoacridine derivatives with benzimidazole or benzoxazole derivatives. Condensation of 2-hydroxy naphthaldehyde with functionalized di

Full text of DOI:10.1016/j.bmc.2006.01.054

Bioorganic & Medicinal Chemistry 14 (2006) 3758–3765
Synthesis, anti-inflammatory, analgesic and kinase (CDK-1,
CDK-5 and GSK-3) inhibition activity evaluation of
benzimidazole/benzoxazole derivatives and some Schiff’s bases
Sham M. Sondhi,a,* Nirupma Singh,a Ashok Kumar,b Olivier Lozachc and Laurent Meijerc
aDepartment of Chemistry, Indian Institute of Technology Roorkee (IIT R), Roorkee-247 667, UA, India
bDepartment of Pharmacology, LLRM Medical College, Meerut, UP, India
cCNRS, Station Biologique, 29680- Roscoff, Bretagne, France
Received 3 December 2005; revised 13 January 2006; accepted 14 January 2006
Available online 9 February 2006
Abstract—A series of N-(acridin-9-yl)-4-(benzo[d]imidazol/oxazol-2-yl) benzamides has been synthesized by the condensation of 9-
aminoacridine derivatives with benzimidazole or benzoxazole derivatives. Condensation of 2-hydroxy naphthaldehyde with func-
tionalized diamines leads to the formation of Schiff’s bases and not imidazole derivatives. All these compounds were characterized
1
by correct FT-IR, H NMR, MS and elemental analyses. These compounds were screened for anti-inflammatory, analgesic and
kinase (CDK-1, CDK-5 and GSK-3) inhibition activities. Compounds 11 and 7e(f) showed good anti-inflammatory (35.8% at
50 mg/kg po) activity and good analgesic activity (60% at 50 mg/kg po), respectively. Compound 3b showed significant in vitro activ-
ity against CDK-5 (IC50 = 4.6 lM) and CDK-1(IC50 = 7.4 lM) and compound 3a showed moderate CDK-5 inhibitory activity
(IC50 = 7.5 lM). The other compounds showed moderate anti-inflammatory and analgesic activities.
ꢀ 2006 Elsevier Ltd. All rights reserved.
1. Introduction
2. Results and discussion
Benzimidazole1–4 and benzoxazole5–7 derivatives pos-
sessing anti-inflammatory, analgesic and kinase inhibi-
tion activities8–10 have been reported in some
literature. Apart from the above-mentioned activities
some interesting efficacy of antiallergic,11 antiprolifera-
tive,12,13 antitumour,14 antiHIV,15 and antibacterial16
activities on benzimidazole derivatives has been report-
ed. Antituberculosis17 activities of benzoxazole deriva-
tives were also reported in the literature.
2.1. Chemistry
N-(Acridin-9-yl)-4-(benzo[d]imidazol-2-yl)benzamide
derivatives 3a–d (Scheme 1) have been synthesized by the
condensation of benzimidazole derivatives 2a–c with 9-
aminoacridine derivatives 1a–b and N-(acridin-9-yl)-4-
(benzo[d]oxazol-2-yl)benzamide derivatives 3e–f (Scheme
1) were synthesized by the reaction of benzoxazole deriv-
ative 2d with 9-aminoacridine derivatives 1a–b by reflux-
ing in methanol or in THF(tetrahydrofuran) for 48 h. The
condensed products 3a–d were purified by chromatogra-
phy on a silica gel column. The precursor benzimidazole
derivatives 2a–c were synthesized by the condensation
of 4-cyanobenzaldehyde with o-phenylenediamines, and
benzoxazole derivative 2d was synthesized by the reaction
of 4-cyanobenzaldehyde with o-amino phenol by reflux-
ing in nitrobenzene.22 The key intermediates, 9-aminoac-
ridines 1a–b, were synthesized by following the reaction
procedure reported in the literature.23 Spectral data and
elemental analyses of compounds 3a–f fully support the
structures assigned to them.
Due to broad spectrum of activities reported in the liter-
ature so far, we have synthesized a number of benzimid-
azole and benzoxazole derivatives targeting for potent
molecules possessing anti-inflammatory, analgesic18–20
and kinase inhibitory activities.21 We will describe in this
paper the synthesis, screening results of in vivo anti-in-
flammatory and analgesic activities and in vitro kinase
inhibitory activities.
Keywords: Benzimidazole derivatives; Benzoxazole derivatives; Schiff’s
bases; Anti-inflammatory; Analgesic; CDK-1; CDK-5 and GSK-3.
*
It is interesting to note that condensation of 9-amino-
acridine derivative 1 (Scheme 1) with benzimidazole or
Corresponding author. Tel.: +91 1332 285811; fax: +91 1332
273650; e-mail: sondifcy@iitr.ernet.in
0968-0896/$ - see front matter ꢀ 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2006.01.054

S. M. Sondhi et al. / Bioorg. Med. Chem. 14 (2006) 3758–3765
3759
R1
R2
N
X
N
NH2
NC
+
R1
R2
O
C
N
X
R3
H
2
N
N
R1
2a Cl
2b NO2
2c CH3 CH3 NH
2d
R2
X
NH
NH
1
H
R3
3
1a R3= H
1b R3= OCH3
H
R1
3a Cl
3b NO2
3c NO2
3d CH3 CH3
3e
3f
R2 R3
X
NH
NH
H
H
O
H
H
H
H
H
MeOH or
THF
;48hrs.
OCH3 NH
H
H
OCH3
NH
O
O
H
H
H
H
R1
R2
NH
N
X
H
N
N C
R3
3'
(crude)
Scheme 1.
benzoxazole derivative 2 should give condensation prod-
uct 30. But we got a hydrolysed product 3 instead of
getting 30 (Scheme 1).
thaldehyde (4) and (un) substituted o-phenylenediamine
(5) were taken in equimolar ratio and then heated
at 140–150 ꢁC in nitrobenzene for 24 h. Solvent was
removed under reduced pressure and the residue left be-
hind was crystallized from THF to give pure condensed
product 7a without affording compound 6 (Scheme 3).
The same reaction was also achieved by using microwave
irradiation as mentioned below.
The formation of 3 can arise from hydrolysis of 30 dur-
ing purification by chromatography on a silica gel col-
umn. A probable mechanism for the formation of 3
from 30 is suggested in Scheme 2. Similar type of obser-
vation is also reported in the literature.24 FT-IR spectra
of compound 3a showed amide stretching at 1707
(NHCO) and FAB-MS of 3a shows MH+ ion peak at
m/z 449 (MH+; 4%) Calcd for C27H17N4OCl m/z 448
and the fragmented cations observed in the mass spectra
fully support the structure of 3a (Scheme 2).
2-Hydroxy naphthaldehyde (4) and o-phenylenedi-
amine (5) were dissolved in methanol in equimolar ra-
tio and then the mixture irradiated in microwave oven
for 30 min at 100 watt. During irradiation, the com-
pound separated out and after irradiation the solvent
was removed under reduced pressure and the crude
product was crystallized from THF to give pure con-
densed product 7a (Scheme 3). The product 7a (Scheme
3) obtained by refluxing in nitrobenzene and by micro-
wave irradiation method was found to be identical
(from m.p., mix m.p.; TLC and co-TLC).
In our earlier studies of this research, the reactions of
p-cyanobenzaldehyde on condensation with o-phenylene-
diamines gave benzimidazole derivatives. We tried to ex-
tend this reaction by using 2-hydroxynaphthaldehyde
and various o-phenylenediamines and 2,3-diamino
pyrimidines. 2-Hydroxy naphthaldehyde (4) was
synthesized from 2-hydroxy naphthol by following the
procedure reported in the literature.25 2-Hydroxy naph-
All other substituted o-phenylenediamines, that is, 5b–e,
4,5-diamino pyrimidine (8) and 3,30-diaminobenzimidine
R1
R2
O
C
R1
R2
NH
C
N
X
N
X
H
H
N
N
N
N
R3
R3
3'
3
(crude)
Silica gel
CHCl3 or CHCl3: EtOAc
Moisture
-NH3
NH3
R1
R2
R1
NH2
C
N
N
H
H
N
N
C
O
N
N
X
X
H
R2
OH2
R3
R3
NH2
C
R1
R2
N
H
N
N
X
OH2
R3
Scheme 2.

3760
S. M. Sondhi et al. / Bioorg. Med. Chem. 14 (2006) 3758–3765
OH
H
N
R1
R1
R2
NH2
NH2
N
NH2
NH2
N
R2
H2N
N
N
OH
HO
OHC
N
6
5
N
8
C
H
MeOH,
M.W*
MeOH,
M.W
H2N
N
R1
R2
OH
9
4
C
H
MeOH,
M.W
H2N
H2N
NH2
NH2
7
10
OH
HO
R1
R2
7a
7b
7c
7d
7e(f)
H
H
H
CH3
H
C
N
N C
COOH
NO2
CH3
Cl(H)
H
H
H2N
NH2
11
H(Cl)
Scheme 3.
(10) were condensed with 2-hydroxy naphthaldehyde (4)
by microwave irradiation to give compounds 7b–e(f), 9
and 11 (Scheme 3).
In diamino pyrimidine (8), –NH2 group in meta position
to both the ring nitrogens is more basic as compared to
the other –NH2 group and reacted with the –CHO
group to give condensation product 9. Similar observa-
tions are already reported in the literature.26
It is interesting to note that the reaction of 5b and 5c
condensed with aldehyde 4 gave only regioisomers 7b
and 7c, respectively. This is due to a more basic amino
group in meta position to –COOH and NO2. Spectral
data of 7b and 7c fully support the structures assigned
to them. In case of 7d, only one product is possible
due to symmetrical structure of 5d.
2.2. Biological activity
The acridinyl-benzimidazole 3a–d, acridinyl-benzoxaz-
ole derivatives 3e–f and Schiff’s bases 7a–e(f) and 11
were tested for: (i) anti-inflammatory activity in the car-
rageenin-induced paw oedema model in albino rats at
50 mg/kg po, according to the method of Winter
et al.27 (ii) analgesic activity28 in the phenylquinone
writhing assay at 50 mg/kg po and (iii) cyclin-dependent
kinase (CDK-1 and CDK-5) and glycogen synthase
kinase (GSK-3) inhibitory activity29–31 and the results
are summarized in Table 1.
4-Chloro-o-phenylenediamine on condensation with 2-
hydroxy naphthaldehyde gave a mixture of two isomeric
products 7e and 7f in the approximate ratio of 4:1. The
ratio can be explained on the basis of inductive and mes-
1
omeric effect of the chloro group. H NMR (300 MHz;
DMSO-d6) of 7e(f) shows doubling of peaks for
–NH2– and –CH@N– at d 5.26 and 5.43 and d 9.61–
9.65, respectively, indicating that condensation product
is a mixture of 7e and 7f. FAB-MS of 7e(f) shows a
MH+ ion peak at m/z 297.92 (35%). Spectral data of
7e(f) fully support the structure assigned to it.
In summary, activity data show that compound 11
exhibited good anti-inflammatory and compound 7e(f)
exhibited good analgesic activity, whereas compound
3b (R1 = NO2, R2 = H, R3 = H and X = NH) showed
Table 1. Anti-inflammatory, analgesic and kinase inhibition activities of compounds 3a–f, 7a–d, e(f) and 11
Compound
Anti-inflammatory activity (%) 50 mg/kg po
Analgesic activity (%) 50 mg/kg po
Kinase IC50 (lM)
CDK-1
CDK-5
GSK-3
3a
3b
0.0
3.5
25
25
0.0
25
25
25
30
20
40
30
60
50
50
15
7.4
7.5
4.6
—
>100
42
—
3c
3d
7.5
0.0
—
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
—
>10
>10
>10
>10
>10
>10
>10
>10
>10
3e
0.0
0.0
3f
7a
26.6
25.5
28.2
28.2
31.4
35.8
38.8
7b
7c
—
—
7d
7e(f)
—
—
11
Ibuprofen
—

S. M. Sondhi et al. / Bioorg. Med. Chem. 14 (2006) 3758–3765
3761
significant activity against CDK-5, while compounds 3a
and 3b showed moderate activity against CDK-5 and
CDK-1, respectively.
Thin-layer chromatography (TLC) was performed on
silica gel G for TLC (Merck) and spots were visual-
ized by iodine vapour or by irradiation with ultravio-
let light (254 nm). Column chromatography was
performed by using Qualigens silica gel for column
chromatography (60–120 mesh). 2% Gum acacia was
used as a vehicle for oral administration of the test
compounds.
Data reported in Table 1 suggest that compound N-
(acridin-9yl)-4-(5-nitro-1H-benzo[d]imidazol-2-yl) (3b)
shows significant in vitro kinase inhibitory activity
against CDK-5 (IC50 4.6 lM) and CDK-1
(IC50 = 7.4 lM), whereas compound 3a has some activ-
ity against CDK-5 (IC50 = 7.6 lM). Compounds 3d–f do
not show any activity towards CDKs and GSK. These
observations revealed that the nature of the substituents
at R1 position of benzimidazole ring of acridinyl-benz-
imidazole derivative has a major impact on CDK inhib-
itory activity. The structures of the active compounds
indicate that substitution at R1 = Cl for 3a and R1 =
NO2 for 3b might enhance potency of hydrogen bond-
ing/acceptor/donor in benzimidazoles 3a and 3b which
probably favours interaction with the active site of the
CDK enzymes, whereas other analogues with
R1 = CH3 and/or H lost their potency.
4.1. General procedure for synthesis of 3
4.1.1. Synthesis of N-(acridin-9-yl)-4-(5-chloro-1H-
benzo[d]imidazol-2yl)benzamide (3a). 4-(5-Chloro-1H-
benzo[d]imidazol-2-yl)benzonitrile
(2a)
(253 mg;
1 mmol) was dissolved in MeOH (30 mL) and to it
was added 9-aminoacridine (1a) (194 mg; 1 mmol). The
reaction contents were heated under reflux for 48 h
and then the solvent was removed under reduced pres-
sure. The solid residue left behind was dissolved in meth-
anol and to this solution was added silica gel (ꢀ20 g)
and then the solvent was removed under reduced pres-
sure to give a crude product absorbed over silica gel.
Column was packed with silica gel using pet. ether as
solvent. Crude product adsorbed over silica gel was
packed on the top of silica gel column and then elution
was done with pet. ether, Chloroform/pet. ether (4:1) to
remove side products. Further elution with chloroform
gave pure condensed product 3a. Similarly compounds
3b–d were synthesized.
In case of Schiff’s bases, compound 4,40bis [1-{(2-amino-
phenylimino) methyl] naphthalen-2-ol (11) showed good
anti-inflammatory activity 35.8% at 50 mg/kg po,
whereas other derivatives showed moderate anti-inflam-
matory activity. The activity in case of 11 increases
because of bis nature which is favourable for anti-in-
flammatory activity, whereas compound [(2-aminophe-
nylimino) methyl] naphthalen-2-ol (7a) showed
significant anti-inflammatory activity. Compounds
7e(f) also showed good anti-inflammatory, probably
due to enhanced activity by the presence of the Cl group.
Compounds 7e(f) and 11 showed good analgesic activi-
ty. Similarly ananlgesic activity of 11 (bis form of 7a)
was increased, as compared to 7a.
Solvent of elution, CHCl3; white solid (0.300 g, 50%);
mp 215 (d) ꢁC; IR (KBr) vmax: 3409, 3279, 1707, 1608,
1545, 1482, 1439, 1297 cmꢁ1 1H NMR (DMSO-d6,
;
300 MHz) d: 7.20–7.30 (m, 4H, 3H Ar + 1H, NH,
exch.); 7.55 (d, 2H, Ar); 7.60 (m, 1H, Ar); 7.75 (m,
3H, Ar); 8.05 (dd, 2H, Ar); 8.25 (dd, 2H, Ar); 8.35 (d,
2H, Ar); 11.75 (s, 1H, NH exch.). FAB MS: m/z 449
Cl
N
O
(MH+, 4%); 255 (
; 35%); 254 (m/z
3. Conclusion
C
N
H
Cl
N
In conclusion, according to the results of screening tests,
the bioactivity of the described compounds shows a
clearcut relation with substitution on R1. This structure
activity relationship observation will be helpful to design
novel analogues of this class of compounds as prospec-
tive kinase inhibitors and anti-inflammatory agents.
255; 100.0%); 228
(
;
2.0%); 227
N
H
Cl
NCO
N
N
(
;12%); 220(
;
2.0%). Anal.
N
H
Calcd for C27H17N4OCl: C, 72.24; H, 3.79; N, 12.48.
Found: C, 72.00; H, 3.82; N, 12.57.
4. Experimental
4.1.2. Synthesis of N-(acridin-9-yl)-4-(5-nitro-1H-
benzo[d]imidazol-2yl)benzamide (3b). Solvent of elution,
pet. ether/EtOAc (4:1); white solid (0.300 g, 50%); mp
215 ꢁC; IR (KBr) vmax: 3294, 1620, 1521, 1473,
Melting points (mp) were determined on a JSGW
apparatus and are uncorrected. Domestic microwave
oven model M197 DL (SAMSUNG) was used for
microwave irradiation. IR spectra were recorded using
1339 cmꢁ1 1H NMR (DMSO-d6 + CDCl3, 300 MHz)
;
1
a Perkin Elmer 1600FT Spectrophotometer. H NMR
d: 7.20–7.23 (d, 2H, Ar); 7.58–7.59 (d, 1H, Ar); 7.68–
7.73 (q, 2H, Ar); 7.84–7.86 (d, 4H, Ar); 7.93–7.96 (d,
1H, Ar); 8.17–8.21 (dd, 1H, Ar); 8.25–8.29 (dd, 1H,
Ar); 8.32 (s, 1H, Ar); 8.39–8.42 (d, 2H, Ar); 8.57 (s,
1H –NH–, exch.). EI MS, do not show M+ ion peak.
spectra were recorded on a Bruker WH-300 Spectrom-
eter in a ca. 5–15% (w/v) solution in DMSO-d6 (TMS
as internal standard). The Mass spectrometer peak
measurements were made by comparison with perflu-
orotributylamine using AEI MS-9 double focusing
O2N
O2N
N
O
N
281 (
1.2%); 239 (
;
C
NH
high-resolution mass spectrometer at
a resolving
N
N
H
H
O
C
power of 15,000. FAB-MS was reordered on Jeol
SX-120 (FAB) spectrometer. Electron impact (EI)
MS were recorded on a Jeol D-300 mass spectrometer.
NCO
N
NH
N
2.8%); 221 (
;1.2%); 220 (
; 1.2%);

3762
S. M. Sondhi et al. / Bioorg. Med. Chem. 14 (2006) 3758–3765
OH
N
N
195 (
; 7.4%); 193 (
1.6%); 136
; 2.2%);179
; 28%);135
(30 mL) and to it was added 9-aminoacridine (1a)
N
H
(194 mg; 1 mmol). The reaction contents were heated
under reflux for 48 h and then the solvent was removed
under reduced pressure to give a crude product. This
crude product was purified by column chromatography
over silica gel. Elution with pet. ether/chloroform (1:1)
removed the side product and further elution with pet.
ether/chloroform (1:4) gave pure product 3e.
O2N
(
;
(
NH
N
O2N
(
N ; 4.5%). Anal. Calcd for C27H17N5O3: C,
70.58; H, 3.70; N, 15.25. Found: C, 70.72; H, 3.83; N,
15.00.
Compound 3f was similarly synthesized.
4.1.3. Synthesis of N-(4-methoxy acridin-9-yl)-4-(5-nitro-
1H-benzo[d]imidazol-2yl)benzamide (3c). Solvent of elu-
tion, pet. ether/EtOAc (1:1); white solid (0.300 g, 65%);
mp >250 ꢁC; IR (KBr) vmax: 3294, 1625, 1585, 1522,
Solvent of elution, pet. ether/CHCl3 (1:4); white solid
(0.300 g, 62%); mp 175 ꢁC; IR (KBr) vmax: 3297, 1702,
1608, 1573, 1492, 1450, 1278 cmꢁ1
;
1H NMR
1072 cmꢁ1 1H NMR (DMSO-d6 + CDCl3, 300 MHz)
;
(CDCl3 + D2O, 300 MHz) d: 7.31–7.36 (t, 4H, Ar);
7.52–7.54 (d, 2H, Ar); 7.71–7.75 (d, 6H, Ar); 8.20–8.30
d: 4.08 (s, 3H, OCH3); 7.13–7.24 (m, 3H, Ar); 7.55–
7.66 (m, 1H, Ar); 7.70–7.73 (d, 1H, Ar); 7.87–7.93 (m,
4H, Ar); 8.14–8.18 (dd, 1H, Ar); 8.27–8.29 (d, 1H,
Ar); 8.39–8.49 (d, 3H, Ar); 8.54 (s, 1H, NH exch.);
10.97 (s, 1H, NH exch.). EI MS (m/z relative intensity)
(t, 4H, Ar). EI MS: No M+ ion peak. 222
O
NH
N
C
N
O
C
(
(
; 2.2%); 221 (
; 14%); 220
; 100%); 195
O
NCO
OH
N
O2N
N
; 100%); 220 (
CN
O
239 (
219 (
(
; 3.0%); 225 (
; 74.6%);
N
N
H
N
OCH3
O
N
OH
OH
NCO
N
(
(
;
0.8%); 194
; 4.1%); 193 (
(
;
4.1%); 194
; 0.6%); 210 (
; 100%); 209
N
N
NH
N
O
N
O
O
; 1.6%); 92 (
;
;
4.7%); 208
(
; 1.1%); 193
N
N
4.2%). Anal. Calcd for C27H17N3O2: C, 78.07; H, 4.09;
N, 10.12. Found: C, 77.82; H, 4.01; N, 10.31.
OCH3
OCH3
N
(
; 1.0%). Anal. Calcd for C28H19N5O4: C,
4.1.6. Synthesis of 4-(benzo[d]oxazol-2-yl)-N-(4-methoxy
acridin-9-yl)benzamide (3f). Solvent of elution, pet.
ether/EtoAc (4:1); white solid (0.300 g, 60%); mp
170 ꢁC; IR (KBr) vmax: 3466, 1610, 1548, 1487, 1445,
N
H
68.71; H, 3.88; N, 14.31. Found: C, 68.93; H, 4.00; N,
14.62.
1279 cmꢁ1 1H NMR (CDCl3, 300 MHz) d: 3.97 (s,
;
4.1.4. Synthesis of N-(acridin-9yl)-4-(5,6-dimethyl-1H-
benzo[d]imidazol-2yl)benzamide (3d). Solvent of elution,
CHCl3/EtOAc (4:1); white solid (0.300 g, 62%); mp
285 ꢁC; IR (KBr) vmax: 3190, 1640, 1611, 1443,
3H, OCH3); 7.39–7.46 (m, 4H, Ar); 7.63–7.66 (q, 2H,
Ar); 7.79–7.86 (m, 5H, 4HAr + 1H, NH, exch.); 8.16–
8.20 (d, 1H, Ar); 8.20–8.43 (m, 4H, Ar). EI MS: No
O
1278 cmꢁ1; H NMR (DMSO-d6, 300 MHz) d: 2.37 (s,
1
NH
C
M+ ion peak: 251
(
;
0.5%); 222
6H, CH3 + CH3); 7.27–7.32 (t, 2H, Ar); 7.45 (s, 2H,
Ar); 7.57–7.60 (d, 2H, Ar); 7.75–7.80 (t, 2H, Ar); 8.02–
8.05 (d, 2H, Ar); 8.26–8.35 (dd, 4H, Ar); 11.80 (s, 1H,
NH exch.). HR MS (m/z; relative intensity). No M+
N
OCH3
O
C
NH
N
O
C
(
;15.3%); 221(
; m/z 251-HCHO;
H3
C
N
O
C
O
ion peak. 249.10197
H3C
(
;
2.32%);
N
H3C
N
H
NCO
N
N
247.11069 (
; 100.00%); 246.10313
CN
15%); 220 (
100%); 219 (
; 100%); 220 (
; 0.5%); 218 (
;
CN
H3C
N
O
H
N
N
(247.11069-; 53.00%); 232.08745 (H C
CN
;
NCO
OH
N
N
3
H
H3C
N
; 0.1%);
58.75%);
221.10695
(
;
2.63%);
H3C
N
H
N
O
OH
195.06837 (
; 9.10%); 178.06426 (
;
N
209 (
; 0.4%); 208 (
; 0.3%); 195
; 5.8%); 92
N
N
N
OCH3
OCH3
2.02%). Anal. Calcd for C29H22N4O: C, 78.73; H, 4.97;
N, 12.66. Found: C, 78.41; H, 5.01; N, 12.51.
N
N
(
; 1.4%); 194 (
O
O
4.1.5. Synthesis of N-(acridin-9-yl)-4-(benzo[d]oxazol-2-
yl)benzamide (3e). 4-(Benzo[d]oxazol-2-yl)benzonitrile
(2d) (220 mg; 1 mmol) was dissolved in MeOH
O
(
; 6.4%). Anal. Calcd for C28H19N3O3: C, 75.50;
H, 4.26; N, 9.43. Found: C, 75.83; H, 4.39; N, 9.71.

S. M. Sondhi et al. / Bioorg. Med. Chem. 14 (2006) 3758–3765
3763
4.2. General procedure for 7
7.32–7.37 (t, 1H, Ar); 7.50–7.56 (m, 1H, Ar); 7.71–7.82
(dd, 2H, Ar); 8.16–8.18 (d, 1H, Ar); 9.39 (s, 1H,
–CH@N–). 15.30 (br s, 1H, –OH, exch.). FAB MS
4.2.1. Synthesis of 1-[(2-aminophenylimino) methyl]
naphthalen-2-ol (7a). 2-Hydroxynaphthaldehyde (4)
(172 mg; 1 mmol) and o-phenylenediamine (5a)
(108 mg; 1 mmol) were taken in methanol (5 mL).
The reaction contents were irradiated in microwave
oven for 30 min at 100-watt power and temperature
of the reaction contents was about 63 ꢁC. Crude prod-
uct so obtained was crystallized from THF to give pure
product 1-[(2-aminophenylimino) methyl] naphthalen-
2-ol (7a).
H2N
CH3
291.68 (MH+; 23%); 290.67 (M+; 28%); 147 (
;
CH3
H2N
CH3
16%); 120
(
;
16%). Anal. Calcd for
HC
N
CH3
C19H18N2O: C, 78.62; H, 6.20; N, 9.65. Found: C,
78.21; H, 6.30; N, 9.90.
4.2.5. Synthesis of 1-[(2-amino-4(5)-chlorophenylimino)
methyl] naphthalen-2-ol (7e(f)). Solvent of crystalliza-
tion, THF; red solid (0.300 g, 60%); mp 230 ꢁC; IR
(KBr) vmax: 3447, 1619, 1478, 1400, 1317 cmꢁ1 1H
;
Similarly were prepared other derivatives, that is, 7b–d
and 7e(f).
NMR (DMSO-d6, 300 MHz) d: 5.26 and 5.43 (2s, 2H,
–NH2 exch.); 6.68–7.15 (m, 3H, Ar); 7.38–7.59 (m, 3H,
Ar); 7.84–7.86 (d, 1H, Ar); 7.96–7.99 (d, 1H, Ar);
8.51–8.60 (q, 1H, Ar); 9.61–9.65 (d,1H, –HC@N–);
15.29–15.35 (d, 1H, OH exch.). Two singlets at 5.26
and 5.43 show existence the of two diastereomers. Also
a doublet for –CH@N– at d 9.61–9.65 indicates the pres-
ence of diastereomers. FAB MS 297.92 (MH+; 32%).
Anal. Calcd for C17H13N2OCl: C, 68.80; H, 4.38; N,
9.44. Found: C, 69.02; H, 4.60; N, 9.21.
Solvent of crystallization, THF; orange solid (0.300 g,
65%); mp 175 ꢁC; IR (KBr) vmax: 3469, 3374, 1609,
1557, 1475, 1400, 1315 cmꢁ1
;
1H NMR (CDCl3,
300 MHz) d: 3.70 (br s, 2H, –NH2; exch.); 6.72–6.85
(m, 2H, Ar); 7.06–7.17 (m, 2H, Ar); 7.28–7.36 (m, 2H,
Ar); 7.47–7.54 (m, 1H, Ar); 7.72–7.82 (m, 2H, Ar);
8.12–8.14 (d, 1H, Ar); 9.40 (s, 1H, –HC@N). 15.05 (br
s, 1H, OH exch.). FAB MS: 263.60 (MH+; 27); 262.60
H2N
; 21%); 92 (H N
; 15%); 91
2
(M+; 22); 119 (
4.2.6. Synthesis of 1-[(4-aminopyrimidin-5-yl-imino)
methyl] naphthalen-2-ol (9). 2-Hydroxynaphthaldehyde
(4) (172 mg; 1 mmol) and 4,5-diaminopyrimidine (8)
(110 mg; 1 mmol) were taken in methanol (5 mL). The
reaction contents were irradiated in microwave oven
for 30 min at 100-watt power and temperature of the
reaction contents was about 63 ꢁC. Crude product so
obtained was crystallized from THF to give pure prod-
uct 1-[(4-aminopyrimidin-5-yl-imino) methyl] naphtha-
len-2-ol (9).
HC
N
NH
(
; 22%). Anal. Calcd for C17H14N2O: C, 77.86;
H, 5.34; N, 10.68. Found: C, 77.62; H, 5.10; N, 10.55.
4.2.2. Synthesis of 4-amino-3-[(2-hydroxynaphthalen-1-
yl)methylene amino)] benzoic acid (7b). Solvent of crys-
tallization, THF; pale yellow solid (0.300 g, 60%); mp
>260 ꢁC; IR (KBr) vmax: 3435, 3334, 1677, 1618, 1400,
1
1290 cmꢁ1; H NMR (DMSO-d6, 300 MHz) d: 5.89 (s,
2H, –NH2 exch.); 6.81–6.84 (d, 1H, Ar); 7.15–7.18 (d,
1H, Ar); 7.37–7.42 (t, 1H, Ar); 7.46–7.66 (m, 2H, Ar);
7.82–7.87 (t, 2H, Ar); 7.97–8.00 (d, 1H, Ar); 8.55–8.58
(d, 1H, Ar); 9.65 (s, 1H, >CH@N–); 15.15 (s, 1H,
–OH, exch.). FAB MS 307.67 (MH+; 65%); 306.70
(M+; 15%); 305.66 (M+ꢁH; 8%). Anal. Calcd for
C18H14N2O3: C, 70.58; H, 4.57; N, 9.15. Found: C,
70.71; H, 4.42; N, 9.29.
Solvent of crystallization, THF; red solid (0.300 g, 86%);
mp 240 ꢁC; IR (KBr) vmax: 3460, 3321, 1653, 1580, 1508,
1398 cmꢁ1 1H NMR (DMSO-d6, 300 MHz) d: 6.98–
;
8.32 (m, 7H, Ar); 8.54–8.57 (d, 1H, Ar); 9.66 (s, 1H,
–CH@N–); 14.29 (s, 1H, OH, exch.); NH2 is expected
under water peak. FAB MS 265.59 (MH+; 32%); 121
H2N
(
N; 33%). Anal. Calcd for C15H12N4O: C, 68.18;
N
HC
N
4.2.3. Synthesis of 1-[(2-amino-5-nitro-phenyl imino)
methyl] naphthalen-2-ol (7c). Solvent of crystallization,
THF; yellow solid (0.300 g, 75%); mp 225 ꢁC; IR
H, 4.54; N, 21.21. Found: C, 68.00; H, 4.61; N, 21.02.
4.2.7. Synthesis of 4,40bis [1-{(2-aminophenylimino)
methyl] naphthalen-2-ol (11). 2-Hydroxynaphthaldehyde
(4) (172 mg; 1 mmol) and 3,30-diaminobenzidine (8)
(214 mg; 1 mmol) were taken in methanol (5 mL). The
reaction contents were irradiated in microwave oven for
30 min at 100-watt power and temperature of the reaction
content was about 63 ꢁC. Crude product so obtained was
crystallized from THF to give pure product 4,40bis[1-{(2-
aminophenylimino) methyl] naphthalen-2-ol (11).
(KBr) vmax: 3469, 3343, 1631, 1484, 1400, 1312 cmꢁ1
;
1H NMR (DMSO-d6 + D2O, 300 MHz) d: 6.86 (d, 1H,
Ar); 7.20 (d, 1H, Ar); 7.42 (t, 1H, Ar); 7.63 (m, 1H,
Ar); 7.81 (d, 1H, Ar); 7.91 (d, 1H, Ar); 8.01 (dd, 1H,
Ar); 8.06 (d, 1H, Ar); 8.30 (d, 1H, Ar); 9.52 (s, 1H,
Ar). FAB MS 308.76 (MH+, 7%). Anal. Calcd for
C17H13N3O3: C, 66.44; H, 4.23; N, 13.68. Found: C,
66.13; H, 4.01; N, 13.80.
4.2.4. Synthesis of 1-[(2-amino-4,5-dimethylphenylimino)
methyl] naphthalen-2-ol (7d). Solvent of crystallization,
THF; red solid (0.300 g, 80%); mp 210 ꢁC; IR (KBr)
vmax: 3444, 3221, 2917, 1615, 1539, 1486, 1400,
Solvent of crystallization, THF; red solid (0.300 g, 85%);
mp 235 ꢁC; IR (KBr) vmax: 3439, 3197, 1607, 1485, 1400,
1312 cmꢁ1 1H NMR (DMSO-d6, 300 MHz) d: 5.26
;
1
1310 cmꢁ1; H NMR (DMSO-d6 + CDCl3, 300 MHz)
(br s, 4H, NH2 + NH2 exch.); 6.94–7.19 (q, 4H, Ar);
7.34–7.89 (m, 12H, Ar); 8.39 (d, 2H, Ar); 9.53–9.61 (d,
2H, 2·–CH@N–). FAB MS 523.73 (MH+; 2%); 522.73
d: 2.21 (s, 6H, CH3 + CH3); 3.40 (br s, 2H, NH2) 6.64
(s, 1H, Ar); 6.96 (s, 1H, Ar); 7.13–7.14 (d, 1H, Ar);

3764
S. M. Sondhi et al. / Bioorg. Med. Chem. 14 (2006) 3758–3765
(M+; 3%). Anal. Calcd for C34H26N4O2: C, 78.16; H,
4.98; N, 10.72. Found: C, 78.41; H, 5.03; N, 10.51.
bovine serum albumin (BSA), nitrophenylphosphate,
leupeptin, aprotinin, pepstatin, soybean trypsin inhibi-
tor, benzamidine and histone H1(type III-S) were
obtained from Sigma Chemicals. [c-33P]ATP was ob-
tained from Amersham. The GS-1 peptide (YRRA
AVPPSPSLSRHSSPHQSpEDEEE) was synthesized by
the Peptide Synthesis Unit, Institute of Biomolecular
Sciences, University of Southampton, Southhampton
SO16 7PX, U.K.
4.3. Anti-inflammatory activity evaluation
Anti-inflammatory activity evaluation27 was carried out
using carrageenin-induced paw oedema in albino rats.
Oedema in one of the hind paws was induced by injec-
tion of carrageenin solution (0.1 mL of 1%) into plan-
tar apponeurosis. The volume of the paw was
measured plethysmographically immediately after and
three hours after the injection of the irritant. The dif-
ference in volume gave the amount of oedema devel-
oped. Percent inhibition of the oedema between the
control group and compound treated groups was calcu-
lated and compared with that of the group receiving a
standard drug.
4.5.2. Buffers
4.5.2.1. Homogenization buffer. 60 mM b-glycerophos-
phate, 15 mM p-nitrophenylphosphate 25 mM Mops
(pH 7.2), 15 mM EGTA, 15 mM MgCl2, 1 mM DTT,
1 mM sodium vanadate, 1 mM NaF, 1 mM phenylphos-
phate, 10 lg leupeptin/mL, 10 lg aprotinin/mL, 10 lg
soybean trypsin inhibitor/mL and 100 lM benzamidine.
4.5.2.2. Buffer A. 10 mM MgCl2, 1 mM EGTA, 1 mM
DTT, 25 mM Tris–HCl, pH 7.5, and 50 lg heparin/mL.
Compounds 3a–f, 7a–e(f) and 11 at 50 mg/kg po sus-
pended in 2% gum acacia exhibited 0.0%, 3.5%, 7.5%,
0.0%, 0.0%, 0.0%, 26.6%, 25.5%, 28.2%, 28.2%, 31.4%
and 35.8%, respectively, anti-inflammatory activity
(Table 1) compared to ibuprofen which showed
38.8% activity at 50 mg/kg po.
4.5.2.3. Buffer C. Homogenization buffer but 5 mM
EGTA, no NaF and no protease inhibitors.
4.5.3. Kinase preparations and assays. Kinase activities
were assayed in Buffer A or C, at 30 ꢁC, at a final
ATP concentration of 15 lM. Blank values were sub-
tracted and activities calculated as pmoles of phosphate
incorporated for 10 min incubation. The activities were
calculated in % of the maximal activity, that is, in the
absence of inhibitors. Controls were performed with
appropriate dilutions of dimethylsulfoxide.
4.4. Analgesic activity evaluation
Analgesia was measured by the writhing assay28 using
Swiss mice (15–20 g). Female mice were screened for
writhing on day
1 by injecting intraperitoneally
0.2 cm3 of 0.02% aqueous solution of phenylquinone.
They were kept on flat surface and the number of
writhes of each mouse was recorded for 20 min. The
mice showing significant writhes (>10) were sorted
out and used for analgesic assay on the following
day. The mice consisting of 5 in each group and show-
ing significant writhing were given orally a 50 mg/kg po
dose suspended in 2% gum acacia of the test com-
pounds 15 min. prior to phenylquinone challenge.
Writhing was again recorded for each mouse in a group
and a percentage protection was calculated using the
following formula.
4.5.3.1. GSK-3a/b. GSK-3a/b was either purified
from porcine brain by affinity purification on an immo-
bilized fragment of axin.29 It was assayed, following a
1/100 dilution in 1 mg BSA/mL, 10 mM DTT, with
5 lL of 40lM GS-1 peptide as a substrate, in buffer A,
in the presence of 15 lM [c-33P]ATP (3000 Ci/mmol;
1 mCi/mL) in a final volume of 30 lL. After 30 min.
incubation at 30 ꢁC, 25 lL aliquots of supernatant were
spotted onto 2.5 · 3 cm pieces of Whatman P81 phos-
phocellulose paper and 20 s later; the filters were washed
five times (for at least 5 min. each time) in a solution of
10 mL phosphoric acid/litre of water. The wet filters
were counted in the presence of 1 mL ACS (Amersham)
scintillation fluid.
Protection¼100ꢁ½fðNo. of writhings for treated miceÞ=
ðNo. of writhings for untreated miceÞgꢂ100ꢃ.
This was taken as a percent of analgesic response and
was averaged in each group of mice. Percent of animals
exhibiting analgesia was determined with each dose.
4.5.3.2. CDK-1/cyclin B. CDK-1/cyclin B was extract-
ed in homogenization buffer from M phase starfish
(Marthasterias glacialis) oocytes and purified by affinity
chromatography on P9CKShs1 Sepharose beads, from
which it was eluted by free P9CKShs1 as reported previ-
ously.30,31 The kinase activity was assayed in buffer C,
with 1 mg histone H1/mL, in the presence of 15 lM
[c-32P]ATP (3000 Ci/mmol; mCi/mL) in a final volume
of 30 lL. After 10 min incubation at 30 ꢁC, 25 lL ali-
quots of supernatant were spotted onto P81 phosphocel-
lulose papers and treated as described above.
Compounds 3a–f, 7a–e(f) and 11 were screened for anal-
gesic activity at 50 mg/kg po. All these compounds
exhibited 25%, 25%, 0.0%, 25%, 25%, 25%, 30%, 20%,
40%, 30%, 60% and 50% analgesic activity, respectively
(Table 1), as compared to ibuprofen which exhibited
50% activity.
4.5. Kinase inhibition activity evaluation29–31
4.5.1. Biochemical reagents. Sodium orthovanadate,
EGTA, EDTA, Mops, b-glycerophosphate, phenyl-
phosphate, sodium fluoride, dithiothreitol (DTT),
4.5.3.3. CDK-5/p25. CDK-5/p25 was reconstituted by
mixing equal amounts of recombinant mammalian
CDK-5 and p25 expressed in Escherichia coli as GST

S. M. Sondhi et al. / Bioorg. Med. Chem. 14 (2006) 3758–3765
3765
11. Nakano, H.; Inoue, T.; Kawasaki, N.; Miyataka, H.;
Matsumoto, H.; Taguchi, T.; Inagaki, N.; Nagai, H.;
Satoh, T. Bioorg. Med. Chem. 2000, 8, 373.
12. Hong, S.-Y.; Chung, K.-H.; You, H.-J.; Choi, I. H.; Chae,
M. J.; Han, J.-Y.; Jung, O.-K.; Kang, S.-J.; Ryu, C.-K.
Bioorg. Med. Chem. Lett. 2004, 14, 3563.
13. Garuti, L.; Roberti, M.; Pizzirani, D.; Pession, A.;
Leoncini, E.; Cenci, V.; Hrelia, S. IL Farmaco 2004, 59,
663.
14. White, A. W.; Curtin, N. J.; Eastman, B. W.; Golding, B.
T.; Hostomsky, Z.; Kyle, S.; Li, J.; Maegley, K. A.;
Skalitzky, D. J.; Webber, S. E.; Yu, X.-H.; Griffin, R. J.
Bioorg. Med. Chem. Lett. 2004, 14, 2433.
15. Novelli, F.; Tasso, B.; Sparatore, F.; Sparatore, A.
Farmaco 1997, 52, 499; Novelli, F.; Tasso, B.; Sparatore,
F.; Sparatore, A. Chem. Abstr. 1998, 128, 238983j.
16. Kazimierczuk, Z.; Andrzejewska, M.; Kaustova, J.;
Klimesova, V. Eur. J. Med. Chem. 2005, 40, 203.
17. Koci, J.; Klimesova, V.; Waisser, K.; Kaustova, J.; Dahse,
H.-M.; Mollmann, U. Bioorg. Med. Chem. Lett. 2002, 12,
3275.
(glutathione-S-transferase) fusion proteins and purified
by affinity chromatography on glutathione–agarose
(vectors kindly provided by Dr. J. H. Wang) (p25 is a
truncated version of p35, the 35KDa CDK-5 activator).
Its activity was assayed in buffer C as described for
CDK-1/cyclin B.
Compounds 3a–f, 7a–e(f) and 11 were screened for
CDK-1, CDK-5 and GSK-3 inhibition activity and
IC50 values are reported in Table 1. Compounds 3a
for CDK-5 (IC50 7.5 lM) and 3b for CDK-1 (IC50
7.4 lM) and CDK-5 (IC50 4.6 lM) exhibited some
interesting inhibition activities.
Acknowledgments
We are thankful to the Head and Mr. Avtar Singh
SAIF-PU, Chandigarh, and the technical staff of the
Chemistry department, I.I.T. Roorkee, Roorkee, for
the spectroscopic studies.
18. Sondhi, S. M.; Singhal, N.; Johar, M.; Reddy, B. S. N.;
Lown, J. W. Curr. Med. Chem. 2002, 9, 1045.
19. Sondhi, S. M.; Singh, N.; Johar, M.; Kumar, A. Bioorg.
Med. Chem. 2005, 13, 6158.
20. Sondhi, S. M.; Goyal, R. N.; Lahoti, A.; Singh, N.;
Shukla, R.; Raghubir, R. Bioorg. Med. Chem. 2005, 13,
3185.
References and notes
21. Sondhi, S. M.; Singh, N.; Lahoti, A. M.; Bajaj, K.;
Kumar, A.; Lozech, O.; Meijer, L. Bioorg. Med. Chem.
2005, 13, 4291.
22. Singhal, N.; Johar, M.; Lown, J. W.; Sondhi, S. M.
Phosphorous, Sulfur Silicon Relat. Elem. 2001, 174,
81.
23. Albert, A.; Ritchie, B. Org. Synth. 1960, 3, 53.
24. Sondhi, S. M.; Singh, N.; Rajvanshi, S.; Johar, M.;
Shukla, R.; Raghubir, R.; Dastdar, S. G. Indian J. Chem.
B 2005, 44, 387.
25. Vogel, A. I. A Text Book of Practical Organic Chemistry;
Longmans: London, 1978, 702.
1. Labanauuskas, L.; Brukstus, A.; Udrenaite, E.; Gaidelis,
P.; Bucinskaite, V. Chemija 2003, 14, 49; Labanauuskas,
L.; Brukstus, A.; Udrenaite, E.; Gaidelis, P.; Bucinskaite,
V. Chem. Abstr. 2003, 139, 180016.
2. Labanauuskas, L. K.; Brukstus, A. B.; Gaidelis, D. G.;
Bucinskaite, V.; Udrenaite, E. B.; Duksas, V. K. Pharm.
Chem. J. 2000, 34, 353; Labanauuskas, L. K.; Brukstus, A.
B.; Gaidelis, D. G.; Bucinskaite, V.; Udrenaite, E. B.;
Duksas, V. K. Chem. Abstr. 2001, 134, 366835.
3. Kataev, V. A.; Khaliullin, A. N., Nasyrov, Kh. M.;
Gailyunas, I. A. Russ. Patent RU 2215002, Chem. Abstr.
2004, 141, 7110. .
26. Sondhi, S. M.; Johar, M.; Rajvanshi, S.; Dastidar, S. G.;
Shukla, R.; Raghubir, R.; Lown, J. W. Aust. J. Chem.
2001, 54, 69.
27. Winter, C. A.; Risley, E. A.; Nuss, G. W. Proc. Soc. Exp.
Biol. Med. 1962, 111, 544.
28. Singh, P. P.; Junnarkar, A. V.; Seshagiri Rao, C.; Verma,
R. K.; Shridhar, D. R. Methods Find. Exp. Clin. Pharma-
col. 1983, 5, 601.
29. Primot, A.; Baratte, B.; Gompel, M.; Borgne, A.; Liabeuf,
S.; Romette, J. L.; Costantini, F.; Meijer, L. Protein Expr.
Purif. 2000, 20, 394.
30. Leclerc, S.; Garnier, M.; Hoessel, R.; Marko, J. A.;
Bibb, J. A.; Snyder, G. L.; Greengard, P.; Biernat, J.;
Mandelkow, E.-M.; Eisenbrand, G. J. Biol. Chem. 2001,
276, 251.
31. Leost, M.; Schultz, C.; Link, A.; Wu, Y.-Z.; Biernat, J.;
Mandelkow, E.-M.; Bibb, J. A.; Snyder, G. L.; Greengard,
P.; Zaharevitz, D. W.; Gussio, R.; Senderovitz, A.;
Sausville, E. A.; Kunick, C.; Meijer, L. Eur. J. Biochem.
2000, 267, 5983.
4. Amin, K.; Dahlstrom, M.; Nordberg, P.; Starke, I. PCT
Int. Appl. WO 97, 47, 603, Chem. Abstr. 1998, 128, 88919.
5. Isomura, Y.; Ito, N.; Homma, H.; Abe, T.; Kubo, K.
Chem. Pharm. Bull. 1983, 31, 3168; Isomura, Y.; Ito, N.;
Homma, H.; Abe, T.; Kubo, K. Chem. Abstr. 1984, 100,
103220.
6. Unelue, S.; Baytas, S. N.; Kupeli, E.; Yesilada, E. Arch.
Pharm. 2003, 336, 310; Unelue, S.; Baytas, S. N.; Kupeli,
E.; Yesilada, E. Chem. Abstr. 2004, 140, 280728.
7. Tsuji, M.; Inoue, T.; Bepu, K.; Saida, M.; Taniguchi, Y.;
Aoki, T.; Furuta, K.; Nakahara, M. Jpn. Kokai, Tokkyo,
Koho JP 63077866, Chem. Abstr. 1984, 109, 128991.
8. Cho, J. M.; Ro, S.; Lee, T. G.; Lee, K. J.; Shin, D.; Hyun,
Y.-Z.; Lee, S. C.; Kim, J. H.; Jeon, Y. H. PCT Int. Appl.
WO 2004065370, Chem. Abstr. 2004, 141, 174171.
9. Clerc, F.; Hamy, F.; Depaty, I.; Angouillant, B.O.;
Roesner, M. Eur Pat. Appl. EP 20030402, Chem. Abstr.
2003, 138, 271683.
10. Bordon, P. F.; Haesslein, J. L. Fr. Demande FR
20040820, Chem. Abstr. 2004, 141, 206968.