Synthesis and preliminary evaluation of some substituted coumarins as anticonvulsant agents

Article abstract of DOI:10.1016/j.bmc.2008.04.021

Some new substituted coumarinylthiazolines, coumarinylthiazolidin-4-ones, and substituted chromenothiazoles were synthesized and evaluated for anticonvulsant activity. Some selected compounds were assayed against seizures induced by pentylenetetrazole (PTZ) and strychnine in mice. Compounds 3b, 6b, and 7b were the most active of the series against PTZ induced seizures. Compound 7b provided anticonvulsant activity (PD₅₀ = 95 mg/kg, ip) at a dose 200 mg/kg compared to phenobarbital (PD₅₀ = 16 mg/kg, ip) at a dose 30 mg/kg (90% protection). No clear correlation was observed between the antiepileptic activity and molecular lipophilicity descriptors of the tested compounds.

Full text of DOI:10.1016/j.bmc.2008.04.021

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry 16 (2008) 5377–5388
Synthesis and preliminary evaluation of some substituted
coumarins as anticonvulsant agents
Kamelia M. Amin, Doaa E. Abdel Rahman* and Yasmin A. Al-Eryani
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Cairo University, Cairo 11562, Egypt
Received 20 December 2007; revised 6 April 2008; accepted 11 April 2008
Available online 15 April 2008
Abstract—Some new substituted coumarinylthiazolines, coumarinylthiazolidin-4-ones, and substituted chromenothiazoles were syn-
thesized and evaluated for anticonvulsant activity. Some selected compounds were assayed against seizures induced by pentylenetet-
razole (PTZ) and strychnine in mice. Compounds 3b, 6b, and 7b were the most active of the series against PTZ induced seizures.
Compound 7b provided anticonvulsant activity (PD50 = 95 mg/kg, ip) at a dose 200 mg/kg compared to phenobarbital
(PD50 = 16 mg/kg, ip) at a dose 30 mg/kg (90% protection). No clear correlation was observed between the antiepileptic activity
and molecular lipophilicity descriptors of the tested compounds.
ꢀ 2008 Elsevier Ltd. All rights reserved.
1. Introduction
A novel class of potential antiepileptics emerged from
studies in the field of plant coumarins. Calophyllolide
and scoparone were found to possess moderate sedative
and anticonvulsant properties.8,9 Recently, it was shown
that both selective MAO-A and MAO-B inhibitors exert
anticonvulsant activity in seizure models. Esuprone,
selective MAO-A inhibitor, exhibited antiepileptic activ-
ity and may be an interesting new approach for treat-
ment of epilepsy.10 Tonabersat, a member of a family
of novel benzoylamino–benzopyran compounds, had
potent anticonvulsant activity in a number of seizure
models with potential for a good therapeutic ratio com-
pared to other anticonvulsants.11
Epilepsy is the most frequent neurologic affection char-
acterized by excessive temporary neuronal discharge.1
The overall prevalence of the disease is 0.5–1.0% of
the population and up to 50 million people world wide.2
Many patients with epilepsy do not respond well to cur-
rently available antiepileptic drugs (AED) such as phe-
nytoin, carbamazepine, diazepam, phenobarbital,
ethusoximide, valproate, valrocemide, vigabatrin, gaba-
pentin, zonisamide, topiramate, tiagabine, felbamate,
retigabine, lamotrigine, and levetiracetam which are
effective towards only 50–80% of the patients and pres-
ent some undesirable side effects such as vertigo, ataxia,
headache, hirsutism, hepato toxicity, gastrointestinal,
and cardiovascular side effects.2–7
In addition many thiourea,12–14 thiazolidinone,15–23
thiazoline,21 and benzothiazole24–27 derivatives exhibit
anticonvulsant activity. In this study, we report the syn-
thesis and the pharmacologic evaluation of new series of
novel synthetic coumarins fitted with functional moieties
believed to enhance antiepileptic activity to assess their
in vivo activity for future development of AED. The new-
ly synthesized compounds belong to the following series,
N-substituted-3-(2-oxo-2H-chromen-6-yl)thioureas 3a–f,
3-substituted-2-(2-oxo-2H-chromen-6-ylimino)thiazoli-
din-4-ones 4a–f, 3-substituted-5-(4-substituted arylid-
ene)-2-(2-oxo-2H-chromen-6-ylimino)thiazolidin-4-ones
5a–e, 3-substituted-4-(4-substituted phenyl)-2-(2-oxo-
2H-chromen-6-ylimino)thiazolines 6a–k, 3-substituted-
4-methyl-2-(2-oxo-2H-chromen-6-ylimino)thiazoline-5-
carboxylic acid ethyl esters 7a–e, 2-substituted amino-7H-
Consequently, a real need exists to develop new anticon-
vulsant compounds to cover seizures which are so far
resistant to presently available drugs. A strategy along
this line is to search for compounds with new modes
of action.
Keywords: Coumarinylthioureas; Coumarinylthiazolidin-4-ones; Cou-
marinylthiazolines; Chromeno[6,5-d]thiazoles; Anticonvulsant.
*
Corresponding author. Tel.: +2 02 23639307; +2 02 24097855; fax:
+2 02 23628426; e-mail: doaaezzat2004@yahoo.com
0968-0896/$ - see front matter ꢀ 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2008.04.021

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K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
chromeno[6,5-d]thiazol-7-ones 8a–c, and 2-(4-oxothiaz-
olidin-2-ylideneamino)-7H-chromeno[6,5-d]thiazol-7-one
11. Some of the newly synthesized compounds were tested
for anticonvulsant activity against chemically induced
convulsion using PTZ and strychnine.
2. Results and discussion
2.1. Chemistry
6-Aminocoumarin 2, was prepared in two steps from cou-
marin following the reported procedures (Scheme 1).28–30
The amine compound 2 was reacted with alkyl/aryl isothi-
ocyanate and gave some new N-substituted-3-(2-oxo-2H-
chromen-6-yl)thioureas 3a–f,Scheme 1. The proposed
structures of compounds 3a–f were confirmed on the basis
of elemental analysis and spectroscopic data (IR, 1H
NMR). The IR spectra showed NH and C@S stretching
bands at 3350–3128 and 1285–1260 cmÀ1, respectively
Scheme 2. Reagents and conditions: (i) ClCH2COOH, CH3COONa,
CH3COOH, reflux, 24–48 h (ii) ArCHO, CH3COONa, CH3COOH,
reflux, 48 h.
1
and the lactone carbonyl at 1720–1707 cmÀ1. The H
NMR spectra of the compounds 3a–f showed the NHs
of the thiourea functionality at the region d = 7.74–
7.85 ppm for the (S@CNHR) and at d = 9.43–
10.01 ppm for the (S@CNHcoumarin). The remaining
protons appeared at the expected chemical shifts.
Cyclization of the thiourea derivatives 3a–f with mono-
chloroacetic acid led to formation of 3-substituted-2-(2-
oxo-2H-chromen-6-ylimino)thiazolidin-4-ones
4a–f,
Scheme 2. The cyclization reaction to 2-iminothiazolid-
inones, possibly proceeds via the non isolated intermedi-
ate
B
following ene-thiolization.31 (Fig. 1) The
corresponding yields of the reaction indicated that com-
pounds 3a–d are more reactive than compounds 3e, f.
This is maybe due to the steric effect of cyclohexyl and
phenyl that reduces the ability of compounds 3e, f to un-
dergo nucleophilic cyclization. The formation of com-
pounds 4a–f was confirmed by the disappearance of
thiourea NHs and C@S absorption bands and appear-
ance of the characteristic lactam CO bands at 1738–
1706 cmÀ1 in the IR spectra. Appearance of a singlet as-
Figure 1. The proposed mechanism of 2-iminothiazolidin-4-one
formation.
signed to methylene thiazolidinone protons at d = 3.67–
4.07 ppm in the H NMR spectra of compounds 4a–f
proved ring closure and confirmed their formation.
1
Condensation of thiazolidinones 4a–c with appropriate
aldehydes in glacial acetic acid buffered with sodium
acetate afforded 5-arylidene-2-imino-4-thiazolidinones
5a–e, Scheme 2. It was noted that the yield of the reac-
tion of 4-nitro or 4-chlorobenzaldehydes with 4a–c was
good and benzaldehyde or its 2-nitro and 4-dimethyl-
amino failed to react with 4a–c. The proposed structures
of 5a–e were confirmed by micro analysis and spectro-
scopic data. Compounds 5a–e exist as potential E and
Z geometrical isomers; the Z conformation of the 5 exo-
cyclic C@C double bond was assigned on the basis of 1H
NMR spectra and on the basis of literature data for ana-
logs 4-thiazolidinones and 2,4-thiazolidindiones.32,33
1
For example, the H NMR spectrum of compound 5a
showed only one kind of the methine proton that
deshielded by the adjacent C@O and was detected at
1
d = 7.41 ppm. Also in H NMR spectrum of 5b, the ab-
sence of the signal of (COCH2) protons of the starting
compounds 4b at d = 4.07 ppm together with the
appearance of methine proton CH at d = 6.49–
8.22 ppm agreed with the proposed structures.
Scheme 1. Reagents and conditions: (i) HNO3, H2SO4, glacial acetic
acid (ii) SnCl2, HCl (iii) RNCS, C2H5OH, reflux, 8 h.

K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
5379
The synthetic steps adopted for the preparation of 3-
substituted-4-(4-substituted phenyl)-2-(2-oxo-2H-chro-
men-6-ylimino)thiazolines 6a–k and 3-substituted-
4-methyl-2-(2-oxo-2H-chromen-6-ylimino)thiazoline-5-
carboxylic acid ethyl esters 7a–e were illustrated in
Scheme 3.
3-Substituted-4-(4-substituted
phenyl)-2-(2-oxo-2H-
chromen-6-ylimino)thiazolines 6a–k were prepared by
refluxing thiourea derivatives 3a–f with 4-bromophena-
cyl bromide or 4-methoxyphenacyl bromide in isopropa-
nol where the products were obtained as hydrobromide
salts. The obtained hydrobromide salts were neutralized
with sodium acetate to get compounds 6a–k. The key
intermediate in the cyclization, enethiol form of the thio-
ureas, is responsible for the formation of the pertinent
thiazolines6a–k according to the following mechanism34
depicted in Figure 2. The structure assignment for the
prepared 6a–k was deduced by elemental and spectral
analysis. The IR spectra of6a–k revealed the disappear-
ance of bands at 3350–3128 cmÀ1 (NHs) and at 1285–
1260 (C@S) and appearance of strong bands at 1610–
1587 (C@N). Furthermore, the 1H NMR spectra of
compounds6a–k lacked the NH signals and showed
new signal at d = 5.59–6.47 ppm attributed to C-5 H
of thiazoline ring.
Figure 2. The proposed mechanism for 2-iminothiazolines formation.
Treatment of unsymmetrical thioureas 3a–e with ethyl
2-chloroacetoacetate in absolute ethanol yielded the
4,5-disubstituted thiazoline derivatives 7a–e via loss of
H2O and HCl on neutralization with ammonia. The
structures of compounds 7a–e were confirmed by ele-
mental analysis and spectroscopic methods. The IR
spectra of these compounds showed disappearance of
NH bands at 3350–3128 cmÀ1 and appearance of a
Scheme 4. Reagents and conditions: (i) Br2, CHCl3, reflux, 30 min.
strong absorption band at 1735–1715 cmÀ1 (C@O of es-
ter) in addition to bands at 1698–1690 cmÀ1 (C@O of
the pyrone ring). The H NMR spectra lacked the NH
1
signals and showed a singlet signal at d = 2.53–
2.75 ppm corresponding to CH3 at 4-position of thiazo-
line ring, quartet signal at d = 4.11–4.23 ppm corre-
sponding to CH2 of ester and triplet signal at
d = 1.12–1.31 ppm corresponding to CH3 of ester.
The synthesis of 2-(substituted amino)-7H-chro-
meno[6,5-d]thiazol-7-ones 8a–c was accomplished as
shown in Scheme 4. Thus when the thioureas 3a–c were
treated with bromine in chloroform, the expected cyc-
lized aminothiazoles 8a–c were obtained. In the IR spec-
tra of 8a–c, the absence of absorption bands at 1285–
1260 cmÀ1 of C@S and appearance of bands 3302–
3217 cmÀ1 (RNH at C-2 thiazole) together with the ab-
1
sence of signal of C-5H and RNHCS protons in the H
NMR spectra of starting compounds 3a at d = 7.74 ppm
agreed with proposed structures.
2-Amino-7H-chromeno[6,5-d]thiazol-7-one 9 was syn-
thesized from 6-aminocoumarin 2 via one pot reaction
as reported by Merchant et al.35 and Mulwad and Shi-
rodkar36 as in Scheme 5. Thus reaction of amine 2 with
potassium thiocyanate and bromine in acetic acid at 0–
5 ꢁC afforded 9.
2-Chloro-N-(7-oxo-7H-chromeno[6,5-d]thiazol-2- yl)acet-
amide/propionamide 10a, b were prepared by refluxing
amine 9 with chloroacetyl chloride or a-chloropropionyl
chloride, respectively, in dry benzene and anhydrous
Scheme 3. Reagents and conditions: (i) Substituted phenacyl bromide,
isopropanol, reflux, 4 h (ii) Ethyl 2-chloroacetoacetate, C2H5OH,
reflux, 24 h.
1
potassium carbonate as catalyst, Scheme 5. H NMR

5380
K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
no-4-thiazolidinone 11 as illustrated in Scheme 5. The
2-thiazolylimino-4-thiazolidinone 11 displays amino–
imino tautomerism. The possibility of the tautomerism
involving a hydroxylic group was excluded by the evi-
dent absence of typical signals of the OH group in IR
1
and H NMR spectra. The IR absorption bands of the
lactam NH group at 3105 cmÀ1 together with strong
bands at 1720 cmÀ1 (C@O) confirms the 11C tautomeric
form.33 (Fig. 3) 1H NMR for compound 11 revealed sin-
glet signal at d = 4.06 ppm corresponding to (CH2) and
singlet signal exchangeable with D2O at d = 12.32 ppm
corresponding to NH.
2.2. Anticonvulsant activity
The brain uptake of drugs is generally related to the
drug lipophilicity, which can be expressed as the log P
and drug ionization constant, pKa.14 The log P of com-
pounds 3b, 3e, 3f, 4b, 5b, 6b, 6d, 6e, 6g, 7a, 7b, 7e, 8a,
10a, and 11 was calculated (Table 11) and the pKa value
of compounds 3b, 6b, and 7b, was calculated. The re-
sulted pKa values of compounds 3b, 6b, and 7b are
5.62 0.42, 7.63 0.33, and 7.44 0.37, respectively.
Scheme 5. Reagents and conditions: (i) Br2, CH3COOH, KSCN (ii)
chloroacylchloride, K2CO3, Benzene, reflux, 24–36 h (iii) NH4SCN,
C2H5OH, reflux, 10 h.
spectrum of compound 10b (R@CH3) showed doublet
signal at d = 1.69 for CH3, quartet signal at d = 4.86 for
CH–Cl and singlet signal at d = 12.94 ppm for NH
(D2O exchangeable). Furthermore, the IR spectrum of
compound 10b revealed the lacking of typical absorption
bands of NH2 and the presence of NH stretching absorp-
tion at 3191 cmÀ1. In addition carbonyl absorption band
of the pyrone ring at 1729 cmÀ1 and that of amide group
at 1697 cmÀ1 were detected.
The selected compounds were screened for anticonvul-
sant activity using PTZ and strychnine induced seizures.
In PTZ test, thioureas 3b, e, and f have shown varying
degrees of activity. It was observed that N-ethyl thiourea
3b showed maximum protection 60% in comparison to
N-cyclohexyl thiourea 3e 30% and N-phenyl thiourea
Heterocyclization of acetamide 10a with potassium thio-
cyanate in refluxing ethanol, efficiently produced 2-imi-
Figure 3. The proposed mechanism of formation and tautomeric structures of 11.

K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
5381
100
90
80
70
60
50
40
30
20
10
0
Compound No.
Figure 4. Bar diagram showing anticonvulsant activity (% protection) of tested compounds and Phenobarbital in PTZ test.
3f 20% derivatives (Table 11, Fig. 4). The lipophilicity
and pKa of the most active one 3b (log P = 2.12 and
pKa = 5.62) were found close to that of the reference
drug phenobarbital (log P = 1.80 and pKa = 7.4).
methoxyphenyl)-2-(2-oxo-2H-chromen-6-ylimino)thiaz-
oline 6b showed more potent activity comparing to the
compound
3-ethyl-4-(4-bromophenyl)-2-(2-oxo-2H-
chromen-6-ylimino)thiazoline 6g (60% and 30%, respec-
tively). Compound 6e with N-phenyl substituent is
slightly more active than compound 6d with N-cyclo-
The thiazolidinone derivative 4b and its 5-(p-nitroben-
zylidene) derivative 5b showed lower activity compared
with the parent thiourea 3b (60%). It was observed that
compound 4b gave 30% protection and introduction of
aryliden group at 5 position of thiazolidinone ring did
not affect the activity (the protection percentage of com-
pound 5b was 30%).
hexyl group in the position
respectively).
3 (30% and 20%,
Interestingly, it was found that the thiazoline-5-carbox-
ylic acid ethyl esters 7b and 7e showed promising anti-
convulsant activity (80% and 60%, respectively)
especially compound 7b that exhibited equipotent activ-
ity at a dose of 200 mg/kg to phenobarbital at a dose of
30 mg/kg (Table 12, Fig. 5) and its LD50 = 1453.78 mg/
kg as calculated from (Table 13). Also it was observed
that the compound 3,4-dimethyl-2-(2-oxo-2H-chro-
Also, it was found that compounds 6b, 6d, 6e, and 6g,
substituted with different iminothiazoline moities at
the sixth position of coumarin showed varying anticon-
vulsant activity, however, compound 3-ethyl-4-(4-
100
90
80
70
60
50
40
30
20
10
3b
6b
7b
Phenobarbital
0
50 mg 100 mg 200 mg 50 mg 100 mg 200 mg
50 mg 100 mg 200 mg 10 mg 20 mg 30 mg
Dose mg/kg
Figure 5. The bar diagram showing anticonvulsant activity (% protection) of compounds 3b, 6b, 7b at doses 50, 100, 200 mg/kg and Phenobarbital at
doses 10, 20, 30 mg/kg in PTZ test.

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K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
men-6-ylimino)thiazoline-5-carboxylic acid ethyl ester
7a devoids anticonvulsant activity (0% protection) while
compound 3-ethyl-4-methyl-2-(2-oxo-2H-chromen-6-
ylimino)thiazoline-5-carboxylic acid ethyl ester 7b gave
Preliminary results of strychnine seizure pattern test re-
vealed that compounds 3e, 3f, 4c, 5d, 6b, 6g, 7b, 8b, and
11 were significantly different from respective strychnine
treated group but not significantly different from respec-
tive phenytoin treated group.
the highest protection.
Also, compounds 8a and 10a that contain benzothiazole
skeleton in their structure devoid any protection against
PTZ while introducing thiazolidinone ring in 10a
showed anticonvulsant activity, for example, compound
11 40% protection.
Briefly, the coumarin iminothiazoline system serves as
prototypic molecule for subsequent molecular modifica-
tion in the search for novel anticonvulsants.
4. Experimental
4.1. Chemistry
Although the convulsant mechanism of action of PTZ is
poorly understood, it is reported that it is able to inhibit
the chloride conductance by binding to sites of GABAA
receptor complex.37
Melting points ꢁC were determined by open capillary
tube method using Electrothermal 9100 melting point
apparatus and were uncorrected. Elemental micro anal-
yses were performed at the micro analytical center, Fac-
ulty of Science, Cairo University and National Research
Center. The IR spectra were recorded as potassium bro-
mide discs on Schimadzu-435 IR spectrophotometer and
For the most significant compounds 3b, 6b, and 7b it
was thought to evaluate these compounds at three
graded doses (50, 100, 200 mg/kg) and compared the re-
sults with that of standard drug phenobarbital at doses
(10, 20, 30 mg/kg). Compound 7b (PD50 = 95 mg/kg) at
a dose of 200 mg/kg was found to be equipotent to phe-
nobarbital (PD50 = 16 mg/kg) at a dose of 30 mg/kg
(90% protection) (Table 12 and Fig. 5).
1
Bruker FT-IR spectrophotometer. The H NMR spec-
tra, in *CDCl3 or DMSO-d6 as a solvent, were recorded
on Varian Gemini 200 spectrophotometer at 200 MHz,
Varian Mercury spectrophotometer at 300 MHz, and
Jeol FX 90Q, 90 MHz. Chemical shifts are reported as
d (ppm) relative to tetramethylsilane (TMS) as internal
standard. Mass spectra were performed on Fennigan
MAT, SSQ 7000 mass spectrophotometer at 70 eV.
The progress of the reaction was determined by thin
layer chromatography, using Macherey–Nagel Alugram
Sil G/UV254 silica gel plates with fluorescent indicator
UV254 and chloroform-ethanol (9.5:0.5) as the eluting
system and the spots were visualized using Vilber Lour-
met ultraviolet lamp at k = 254 nm.
In the model of convulsion induced by strychnine (Table
14), all compounds being tested except compounds 3b,
5b, 6d, 7e, and 10a significantly increased the average
survival time in this model but less than that activity ob-
served with phenytoin drug.
It is known that strychnine directly antagonize the
inhibitory spinal reflexes of glycine.37 All the com-
pounds that increase the survival time therefore, might
cause seizure suppression by acting on glycine inhibitory
mechanisms.
4.1.1. General procedure for synthesis of N-substituted-3-
(2-oxo-2H-chromen-6-yl)thioureas 3a–f. (Scheme 1,
Tables 1, 2). A mixture of 2 (1.61 g, 10 mmol) and or-
ganic isothiocyanate (10 mmol) in absolute ethanol
(25 mL) was heated under reflux for 8 h. The crude
product was collected, washed with water, dried, and
crystallized from suitable solvent.
3. Conclusion
Although the preliminary results revealed only marginal
activity of the tested compounds, it is apparent from the
data of PTZ test that:
• Coumarinylthiourea and coumarinylthiazoline with
N-ethyl substituent showed more protection than
the compounds having N-cyclohexyl or N-phenyl
substituents.
• Cyclization of thiourea derivatives to thiazolidinone
decreases the activity, while cyclization to thiazoline
increases the activity.
• Substitution of the thiazoline derivatives at 4 and 5
positions with methyl and carboxylic acid ethyl ester,
respectively, highly increases the activity.
• The tricyclic derivatives with free amino group at
position 2 of thiazole ring devoid activity while intro-
ducing the thiazolidinone ring markedly increases the
activity.
• Compound 7b was found to be the most active com-
pound, it devoid hypnotic and sedation and has LD50
equal to 1453.78 mg/kg which is higher than that of
phenobarbital.
4.1.2. General procedure for synthesis of 3-substituted-2-
(2-oxo-2H-chromen-6-ylimino)thiazolidin-4-ones
4a–f.
(Scheme 2, Tables 3, 4). A mixture of 3a–f (10 mmol),
monochloroacetic acid (0.95 g, 10 mmol), and anhy-
drous sodium acetate (2.0 g, 25 mmol) in glacial acetic
acid (30 mL) was refluxed for 24–48 h. After cooling,
the reaction mixture was poured onto ice-cold water.
The precipitate was filtered, washed with water, dried,
and crystallized from ethanol.
4.1.3. General procedure for synthesis of 3-substituted-5-
(4-substituted benzylidene)-2-(2-oxo-2H-chromen-6-yli-
mino)thiazolidin-4-ones 5a–e (Scheme 2, Tables 5, 6).
To a well stirred solution of 4a–c (4 mmol) in acetic
acid (35 mL) buffered with sodium acetate (1.0 g,
12 mmol), the appropriate aryl aldehyde (6 mmol)
was added. The solution was refluxed for 48 h then

K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
Table 1. Physical and analytical data of compounds 3a–f
5383
Mol. formula
Mol. wt.
Yield %
MP (ꢁC)
Microanalysis Calc. (Found)
C
H
N
a
b
c
d
e
f
C
C
11H10N2O2S
12H12N2O2S
234.28
248.31
262.33
260.32
302.40
348.81
c
84c
77a
80a
64a
76b
58d
240–242
211–213
190–193
156–159
233–234
241–245
56.40 (55. 95)
58. 05 (58.40)
59.52 (60. 01)
59.98 (60.00)
63.55 (63. 56)
55.10 (55.50)
4.30 (4.86)
4.87 (5.20)
5.38 (5.10)
4.65 (4.70)
6.00 (6.57)
3.76 (3.33)
11.96 (11.39)
11.28 (11.45)
10.68 (10.37)
10.76 (10.77)
9.26 (9.21)
C13H14N2O2S
C13H12N2O2S
C16H18N2O2S
C
16H11ClN2O2SÆH2O
8.03 (8.05)
Crystallization solvent: aEthanol, bCHCl3, DMF/ethanol, dDMF/H2O.
Table 2. IR and 1H NMR spectral data of compounds 3a–f
IR (c, cmÀ1
)
1H NMR (d, ppm)
aj 3350, 3140(2NH), 3000(CHarom), 2950(CH3), 1720(CO), 1620,
1570, 1520(NH, C@C), 1260(C@S)
2.94(s, 3H, CH3), 6.50(d, 1H, C-3H), 7.38(d, 1H, C-7H), 7.57(d,
1H, C-8H), 7.74(s, 2H, C-5H, NH), 8.07(d, 1H, C-4H), 9.65(s, 1H,
NH)
b
c
3334, 3139(2NH), 3070(CHarom), 2977(CH3, CH2), 1712(CO),
1620, 1545, 1520(NH, C@C), 1277(C@S)
1.14(t, 3H, CH3), 3.42(q, 2H, CH2CH3), 6.42–8.10(m, 6H,
aromaticH, NH), 9.5(s, 1H, NH, exchanged with D2O)
3329, 3139(2NH), 2929, 2861(CH3, CH2), 1712 (CO), 1620, 1552,
1518(NH, C@C), 1285(C@S)
0.91(t, 3H, CH3), 1.46–1.62(m, 2H, CH2CH2CH3), 3.68(t, 2H,
CH2CH2CH3), 6.44–8.11(m, 6H, aromaticH, NH), 9.56(s, 1H, NH
exchanged with D2O)
d
e
f
3331, 3128(2NH), 2982(CH2, CH), 1708(CO), 1620, 1544(NH,
C@C), 1284(C@S)
4.12(d, 2H, CH2CH@CH2), 5.17(t, 2H, CH2CH@CH2), 5.78–
5.89(m, 1H, CH2CH@CH2), 6.45–8.12(m, 6H, aromaticH,NH),
9.65(s, 1H, NH exchanged with D2O)
3314, 3132(2NH), 3051(CHarom), 2972, 2928, 2851(CH2, CH),
1713(CO), 1620, 1544, 1509(NH, C@C), 1278(C@S)
1.21–1.95(m, 10H, cyclohexylH except C-1H), 4.09(m, 1H,
cyclohexylC-1H), 6.47–8.09(m, 6H, aromaticH, NH), 9.43(s, 1H,
NH)
3261(2NH), 3040(CHarom), 1707(CO), 1619, 1566, 1527(NH,
C@C), 1283(C@S)
6.51–8.14(m, 9H, aromaticH), 10.01(s, 2H, 2NH, exchanged with
D2O)
j MS: m/z (%) 234, M+, (99.39).
Table 3. Physical and analytical data of compounds 4a–f
Mol. formula
Mol. wt.
Yield %
MP (ꢁC)
Microanalysis Calc. (Found)
C
H
N
a
b
c
d
e
f
C
13H10N2O3S
C14H12N2O3S
15H14N2O3S
C15H12N2O3S
274.30
288.33
302.35
300.34
342.42
370.82
76
81
70
79
43
30
172–175
154–157
72–74
56.92 (57.22)
58.32 (58.30)
59.59 (59.94)
59.99 (60.00)
63.14 (63.15)
58.30 (58.69)
3.67 (3.91)
4.19 (4.00)
4.67 (5.00)
4.03 (4.10)
5.30 (5.34)
2.99 (3.09)
10.21 (10.48)
9.72 (9.47)
9.27 (8.85)
9.33 (9.33)
8.18 (7.93)
7.55 (7.46)
C
83–87
210–212
87–90
C18H18N2O3S
C18H11ClN2O3S
cooled to room temp. The separated solid was filtered,
washed with water, dried and crystallized from
ethanol.
4.1.5. General procedure for synthesis of 3-substituted-4-
methyl-2-(2-oxo-2H-chromen-6-ylimino)thiazoline-5-car-
boxylic acid ethyl esters 7a–e (Scheme 3, Tables 9, 10).
The appropriate substituted thioureas 3a–e (1 mmol)
were dissolved in absolute ethanol (10 mL) and ethyl
2-chloroacetoacetate (0.16 g, 1 mmol) was added. The
reaction mixture was refluxed for 24 h. The liquid was
chilled and treated with 3 N ammonium hydroxide to
pH 8 then diluted with water (30 mL). The precipitate
formed was filtered, washed with water, dried, and crys-
tallized from ethanol.
4.1.4. General procedure for synthesis of 3-substituted-4-
(4-substituted
phenyl)-2-(2-oxo-2H-chromen-6-ylimi-
no)thiazolines 6a–k (Scheme 3, Tables 7, 8). A mixture
of 3a–f (2 mmol), 4-substituted phenacyl bromide
(2.5 mmol) in isopropanol (20 mL) was refluxed for
4 h, the solvent was filtered and the residue was tritu-
rated with 10% sodium acetate. The solution obtained
was extracted several times with chloroform and the
combined organic extracts were washed with water,
dried, and evaporated. The solid formed was collected
and crystallized from ethanol.
4.1.6. General procedure for synthesis of 2-substituted
amino-7H-chromeno[6,5-d]thiazol-7-one 8a–c (Scheme 4).
To a suspension of substituted thioureas 3a–c (4 mmol)

5384
Table 4. IR and 1H NMR spectral data of compounds 4a–f
IR (c, cmÀ1
K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
)
1H NMR (d, ppm)
aj
bjj
c
3053(CHarom.), 2991, 2934(CH3, CH2), 1731, 1706(2CO),
1630(C@N), 1562(C@C)
*3.32(s, 3H, CH3), 3.86(s, 2H, COCH2), 6.42–7.69(m, 5H,
aromaticH)
3100(CHarom.), 2980, 2920(CH3, CH2), 1738, 1720(2CO),
1620(C@N), 1560(C@C)
1.23(t, 3H, CH3), 3.8(q, 2H, CH2CH3), 4.07(s, 2H, COCH2), 6.49–
8.12(m, 5H, aromaticH)
3091(CHarom.), 2971, 2934, 2878(CH3, CH2), 1734(2CO),
1618(C@N), 1563(C@C)
0.83(t, 3H, CH3), 0.99–1.07(m, 2H, CH2CH2CH3), 3.72(t, 2H,
CH2CH2CH3), 4.07(s, 2H, COCH2), 6.46–8.10(m, 5H, aromaticH)
djjj 3070(CHarom.), 2990, 2930(CH2, CH), 1735, 1710(2CO),
1640(C@N), 1560(C@C)
*3.87(s, 2H, COCH2), 4.46(d, 2H, CH2CH@ CH2), 5.28(t, 2H,
CH2CH@CH2), 5.87–5.92(m, 1H, CH2CH@CH2), 6.42–7.69(m,
5H, aromaticH)
e
3059(CHarom.), 2928, 2853(CH2), 1728 (2CO), 1628(C@N),
1563(C@C)
*1.16–2.34(m, 10H, cyclohexylH except C-1H), 3.67(s, 2H,
COCH2), 4.29–4.45(m, 1H, cyclohexyl C-1H), 6.32–7.61(m, 5H,
aromaticH)
*
f
3061(CHarom.), 2972, 2927(CH2), 1728 (2CO), 1630 (C@N),
1565(C@C)
4.03(s, 2H, COCH2), 6.41–7.75(m, 9H, aromaticH)
j MS: m/z (%) 274, M+, (62.37).
jj MS: m/z (%) 288, M+, (100).
jjj MS: m/z (%) 300, M+, (100).
Table 5. Physical and analytical data of compounds 5a–e
Mol. formula
Mol. wt.
Yield %
MP (ꢁC)
Microanalysis Calc. (Found)
C
H
N
a
b
c
C
20H13N3O5S
C21H15N3O5S
22H17N3O5S
C21H15ClN2O3S
22H17ClN2O3S
407.41
421.43
435.46
410.88
424.91
33
34
34
29
70
272–275
234–236
232–234
189–192
177–179
58.96 (59.09)
59.85 (59.81)
60.68 (60.84)
61.39 (61.45)
62.19 (62.18)
3.22 (3.21)
3.59 (3.21)
3.93 (4.08)
3.68 (3.50)
4.03 (4.15)
10.31 (10.21)
9.97 (9.95)
9.65 (9.68)
6.82 (6.78)
6.59 (6.91)
C
d
e
C
Table 6. IR and 1H NMR spectral data of compounds 5a–e
IR (c, cmÀ1
)
1H NMR (d, ppm)
a
3073(CHarom), 2922, 2850(CH3, CH), 1727, 1644(2CO),
1609(C@N), 1562(C@C), 1521, 1344(NO2)
3.36(s, 3H, CH3), 6.56–8.33(m, 10H, aromaticH, @CH)
bj 3063(CHarom.), 2927, 2852(CH3, CH2, CH), 1727, 1641 (2CO),
1610(C@N), 1562(C@C), 1520, 1340(NO2)
1.3(t, 3H, CH3), 3.49(q, 2H, CH2CH3), 6.49–8.22(m, 10H,
aromaticH, @CH)
c
d
e
3064(CHarom.), 2933, 2873(CH3, CH2, CH), 1745, 1709 (2CO),
1646(C@N), 1609, 1561 (C@C), 1514, 1338(NO2)
0.95(t, 3H, CH3), 1.68–1.92(m, 2H, CH2CH2CH3), 3.23 (t, 2H,
CH2CH2CH3), 6.55–8.19(m, 10H, aromatic H, @CH)
2950(CH3, CH2, CH), 1725, 1640(2CO), 1610(C@N), 1590,
1560(C@C).
*1.31(t, 3H, CH3), 4.0(q, 2H, CH2CH3), 6.39–7.65(m, 10H,
aromatic H, @CH)
3060(CHarom.), 2961, 2929, 2875, 2851(CH3, CH2, CH), 1729,
1637(2CO), 1607(C@N), 1562(C@C)
0.94(t, 3H, CH3), 3.00–3.18(m, 2H, CH2CH2CH3), 3.63 (t, 2H,
CH2CH2CH3), 6.59–7.76(m, 10H, aromaticH, @CH)
j MS: m/z (%) 422, M++1, (13.64).
in chloroform (10 mL), bromine (4 mmol) in chloroform
(5 mL) was added dropwise with stirring. The mixture
was refluxed in a water bath for 30 min then solvent
was evaporated. The product was treated with hydro-
chloric acid, filtered and the filtrate neutralized with
ammonia. The precipitate obtained was filtered, dried
and crystallized from ethanol.
CH3), 6.54 (d, 1H, C-3H), 7.3 (d, 1H, C-7H), 7.64 (d,
1H, C-8H), 8.08 (d, 1H, C-4H), 8.15–8.16 (m, 1H, NH
exchanged with D2O). MS: m/z (%) 232, M+, (33.61).
Anal. Found: C 56.66, H 3.74, N 12.02. For
C11H8N2O2S (232.26): C 56.88, H 3.47, N 12.06.
4.1.6.2. 2-Ethylamino-7H-chromeno[6,5-d]thiazol-7-
one 8b. Yield 54%, mp 219–222 ꢁC, IR: 3292 (NH),
3022 (CHarom.), 2967, 2922, 2865 (CH3, CH2), 1713
4.1.6.1. 2-Methylamino-7H-chromeno[6,5-d]thiazol-7-
one 8a. Yield 48%, mp 280–283 ꢁC, IR: 3302 (NH), 3013
(CHarom.), 2938, 2893 (CH3), 1685 (CO), 1615 (C@N),
1593, 1551 (NH, C@C) cmÀ1. 1H NMR: d = 2.99 (d, 3H,
1
(CO), 1617 (C@N), 1590, 1553 (NH, C@C) cmÀ1. H
NMR: d = 1.21 (t, 3H, CH3), 3.41 (q, 2H, CH2CH3),
6.49 (d, 1H, C-3H), 7.25 (d, 1H, C-7H), 7.58 (d, 1H,

K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
Table 7. Physical and analytical data of compounds 6a–k
5385
Mol. formula
Mol. wt.
Yield %
MP (ꢁC)
Microanalysis Calc. (Found)
H
C
N
a
b
c
d
e
f
C
C
20H16N2O3S
21H18N2O3S
364.43
378.45
392.48
432.54
460.94
413.30
427.32
441.35
439.33
481.41
509.81
66
98
66
75
25
70
88
68
98
70
60
117–120
149–152
117–120
229–231
120–122
218–223
201–205
172–176
196–199
200–201
197–199
65.92 (66.34)
66.65 (67.10)
67.33 (66.83)
69.42 (69.50)
65.14 (65.34)
55.22 (55.74)
56.22 (55.85)
57.15 (57.40)
57.41 (57.71)
59.88 (60.43)
56.54 (56.80)
4.43 (4.08)
4.79 (4.59)
5.14 (5.44)
5.59 (5.71)
3.72 (4.00)
3.17 (3.08)
3.54 (4.10)
3.88 (3.88)
3.44 (3.48)
4.40 (4.4 6)
2.77 (3.00)
7.69 (7.51)
7.40 (7.20)
7.14 (7.05)
6.48 (6.17)
6.08 (6.10)
6.78 (6.48)
6.56 (6.75)
6.35 (6.63)
6.38 (6.67)
5.82 (6.12)
5.49 (5.45)
C22H20N2O3S
C25H24N2O3S
C25H17ClN2O3S
C
19H13BrN2O2S
g
h
i
C20H15BrN2O2S
C21H17BrN2O2S
C21H15BrN2O2S
j
C24H21BrN2O2S
C24H14BrClN2O2S
k
Table 8. IR and 1H NMR spectral data of compounds 6a–k
IR (c, cmÀ1
)
1H NMR (d, ppm)
a
b
c
3066(CHarom.), 2949, 2925, 2829(CH3, CH), 1711(CO),
1593(C@N), 1555(C@C)
3.30(s, 3H, NCH3), 3.82(s, 3H, OCH3), 6.16(s, 1H, thiazoline C-
5H), 6.40–7.93 (m, 9H, aromaticH)
3104(CHarom.), 2966, 2926, 2833(CH3, CH2, CH), 1711(CO),
1595(C@N), 1553(C@C)
*1.20(t, 3H, CH3), 3.82–3.88(m, 5H, CH2CH3, OCH3), 5.71(s, 1H,
thiazoline C-5H), 6.36–7.66(m, 9H, aromaticH)
3031(CHarom.), 2960, 2874, 2840(CH3, CH2, CH), 1708(CO),
1598(C@N), 1558 (C@C)
0.71(t, 3H, CH3), 1.55–1.8(m, 2H, CH2CH2CH3), 3.39(t, 2H,
CH2CH2CH3), 3.83(s, 3H, OCH3), 6.12(s, 1H, thiazoline C-5H),
6.52–8.0(m, 9H, aromaticH).
d
e
3001(CHarom.), 2930, 2856(CH3, CH2, CH), 1724(CO),
1598(C@N), 1558(C@C)
*1.06–2.61(m, 10H, cyclohexylH except C-1H), 3.81(s, 3H, OCH3),
3.90–3.97(m, 1H, cyclohexyl C-1H), 5.59(s, 1H, thiazoline C-5H),
6.31–7. 62(m, 9H, aromaticH).
3066(CHarom.), 2920, 2846(CH3, CH), 1727 (CO), 1601(C@N),
1559(C@C)
3.72(s, 3H, OCH3), 6.47(s, 1H, thiazoline C-5H), 6.52–8.06(m,
13H, aromaticH).
f
2913(CH3, CH), 1712(CO), 1587(C@N), 1558 (C@C)
3100(CHarom.), 1710(CO), 1610(C@N), 1590, 1560(C@C)
3.7(s, 3H, CH3), 6.25–8.07(m, 10H, aromaticH, thiazoline C-5H)
gj
*1.22 (t, 3H, CH3), 3.87(q, 2H, CH2CH3), 5.79(s, 1H, thiazoline C-
5H), 6.39–7.7(m, 9H, aromaticH).
h
i
3027(CHarom.), 2962, 2933, 2873(CH), 1710(CO), 1608(C@N),
1586, 1558 (C@C)
0.72(t, 3H, CH3), 1.42–1.78(m, 2H, CH2CH2CH3), 3.74(t, 2H,
CH2CH2CH3), 6.24(s, IH, thiazoline C-5H), 6.42–8.08(m, 9H,
aromaticH).
3030(CHarom.), 1710(CO), 1610(C@N), 1590(C@C)
4.42(d, 2H, CH2CH@CH2), 4.92(d, 1H, CH2CH@ CHaHb), 5.11(d,
1H, CH2CH@CHaHb), 5.85–5.90(m, 1H, CH2CH@CH2), 6.33(s,
1H, thiazoline C-5H), 6.44–8.04(m, 9H, aromaticH).
j
3097(CHarom.), 2928, 2848(CH2, CH), 1731 (CO), 1607(C@N),
1559 (C@C)
*1.04–2.62(m, 10H, cyclohexylH except C-1H), 3.60–3.72(m, 1H,
cyclohexyl C-1H), 5.61(s, 1H, thiazoline C-5H), 6.3–7.61(m, 9H,
aromaticH).
kjj 3080(CHarom.), 2920(CH), 1725(CO), 1610 (C@N), 1580(C@C)
6.4–8.0(m, 14H, aromaticH, thiazoline C-5H).
j MS: m/z (%) 426, M+, (27.89) and 428, M++2, (29.94).
jj MS: m/z (%) 508, M+, (18.34) and 510, M++2, (24.23).
Table 9. Physical and analytical data of compounds 7a–e
Mol. formula
Mol. wt.
Yield %
MP (ꢁC)
Microanalysis Calc. (Found)
C
H
N
a
b
c
d
e
C
17H16N2O4S
344.39
358.42
372.45
370.43
412.51
69
93
88
89
31
150–152
146–150
96–100
59.29 (59.46)
60.32 (60.37)
61.27 (61.50)
61.61 (61.85)
64.06 (64.21)
4.68 (4.82)
5.06 (4.75)
5.41 (5.66)
4.90 (5.00)
5.86 (5.74)
8.13 (8.10)
7.82 (7.76)
7.52 (7.50)
7.56 (7.51)
6.79 (6.79)
C18H18N2O4S
C19H20N2O4S
C19H18N2O4S
121–122
168–170
C
22H24N2O4S

5386
Table 10. IR and 1H NMR spectral data of compounds 7a–e
IR (c, cmÀ1
K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
)
1H NMR (d, ppm)
aj 3065(CHarom.), 2981, 2921, 2851(CH3, CH2), 1735, 1697(2CO),
1617(C@N), 1562 (C@C)
*1.29(t, 3H, COOCH2CH3), 2.61(s, 3H, CH3), 3.5(s, 3H, NCH3),
4.23 (q, 2H, COOCH2CH3), 6.4–7.69(m, 5H, aromatic H)
b
c
d
2981, 2917, 2852(CH3, CH2), 1717, 1690 (2CO), 1590 (C@N),
1558(C@C)
*1.15–1.31(m, 6H, NCH2CH3, COOCH2CH3), 2.53(s, 3H, CH3),
3.95(q, 2H, NCH2CH3), 4.13(q, 2H, COOCH2CH3), 6.29–7.61(m,
5H, aromatic H)
3046(CHarom.), 2965, 2933, 2875(CH3, CH2), 1718, 1692(2CO),
1600 (C@N), 1561(C@C)
1.12–1.28(m, 8H, NCH2CH2CH3, COOCH2CH3), 2.61(s, 3H,
CH3), 3.67(q, 2H, NCH2CH2CH3), 4.16(q, 2H, COOCH2CH3),
6.42–7.61(m, 5H, aromaticH)
3056(CHarom.), 2979, 2925, 2852(CH3, CH2, CH), 1715(2CO),
1599(C@N), 1560 (C@C)
1.21(t, 3H, COOCH2CH3), 2.75(s, 3H, CH3), 4.17(q, 2H,
COOCH2CH3), 4.7(d, 2H, CH2CH@CH2), 5.2(t, 2H, CH2CH@
CH2), 5.8–6.2(m, 1H, CH2CH@CH2), 6.43–8.10(m, 5H, aromatic
H)
e
3078(CHarom.), 2921, 2852(CH3, CH2, CH), 1721, 1698(2CO),
1592 (C@N), 1562 (C@C)
1.17–1.82(m, 13H, cyclohexyl H except C-1H, COOCH2CH3), 2.66
(s, 3H, CH3), 4.11–4.22(m, 3H, COOCH2CH3, cyclohexyl C-1H),
6.5–8.1 (m, 5H, aromaticH)
j MS: m/z (%) 344, M+, (47.87).
C-8H), 8.02 (d, 1H, C-4H), 8.16 (t, 1H, NH exchanged
with D2O). Anal. Found: C 58.44, H 4.26, N 11.35.
For C12H10N2O2S (246.29): C 58.52, H 4.09, N 11.37.
D2O). Anal. Found: C 50.88, H 3.54, N 8.95. For
C13H9ClN2O3S (308.75): C 50.57, H 2.94, N 9.07.
4.1.8. Synthesis of 2-(4-oxothiazolidin-2-ylideneamino)-
7H-chromeno[6,5-d]thiazol-7-one 11. (Scheme 5). A mix-
ture of 10a (0.29 g, 1 mmol) and ammonium thiocyanate
(0.15 g, 2 mmol) in absolute ethanol (50 mL) was re-
fluxed for 10 h, filtered, washed with water, and crystal-
lized from ethanol. Yield 93%, mp >300 ꢁC, IR: 3105
(NH), 3049 (CHarom.), 2917, 2808 (CH2), 1720
4.1.6.3. 2-Propylamino-7H-chromeno[6,5-d]thiazol-7-
one 8c. Yield 75%, mp 135–139 ꢁC, IR: 3217 (NH),
3080 (CHarom.), 2961, 2908 (CH3, CH2), 1725 (CO),
1628 (C@N), 1556 (NH, C@C) cmÀ1
.
1H NMR:
d = 0.95 (t, 3H, CH3), 1.56–1.90 (m, 2H, CH2CH2CH3),
3.62 (t, 2H, CH2CH2CH3), 6.40–8.2 (m, 5H, aromaticH,
NH exchanged with D2O). Anal. Found: C 60.05, H
4.60, N 10.75. For C13H12N2O2S (260.32): C 59.98, H
4.65, N 10.76.
(2CO), 1618 (C@N), 1575 (C@C) cmÀ1 1H NMR:
.
d = 4.06 (s, 2H, CH2), 6.55 (d, 1H, C-3H), 7.42 (d, 1H,
C-7H), 7.90 (d, 1H, C-8H), 8.12 (d, 1H, C-4H), 12.32
(t, 1H, NH exchanged with D2O). Anal. Found: C
49.19, H 2.44, N 13.21. For C13H7N3O3S2 (317.35): C
49.20, H 2.22, N 13.24.
4.1.7. General procedure for synthesis of 2-chloro-N-(7-
acetamide/propi-
oxo-7H-chromeno[6,5-d]thiazol-2-yl)
onamide 10a, b. (Scheme 5). A mixture of 9 (0.33 g,
1.5 mmol), the corresponding chloroacylchloride
(1.5 mmol), and potassium carbonate (0.41 g, 3 mmol)
in dry benzene (20 mL) was refluxed for 24–36 h. The
hot solution was filtered, the solvent was evaporated un-
der reduced pressure and the residue was crystallized
from the appropriate solvent.
4.2. Anticonvulsant Screening
Eighteen selected compounds from the target com-
pounds 3–8, 10, and 11 were evaluated for anticonvul-
sant activity against PTZ and strychnine induced
seizures in mice.
4.1.7.1. 2-Chloro-N-(7-oxo-7H-chromeno[6,5-d]thia-
zol-2-yl)acetamide 10a. Yield 90%, crystallized from
CHCl3, mp 257–260 ꢁC, IR: 3170 (NH), 3070 (CHa-
rom.), 2988, 2939 (CH2), 1735, 1659 (2CO), 1622
4.2.1. PTZ seizure pattern test. Male albino mice weigh-
ing 25–30 g were housed in groups of 10. The animals
were acclimated to their environment for at least 2 days
before the experiments and were allowed free access to
food and water before being tested. The tested com-
pounds were suspended in water and 2% Tween 80
and administered to animals at a dose of 100 mg/kg ip.
1 h after the administration of the tested compound,
mice were injected PTZ (112 mg/kg) ip. Animals devoid
of generalized convulsions were considered to be pro-
tected and results were represented as percentage protec-
tion.38 Standard drug used was phenobarbital sodium at
the dose of 30 mg/kg (Table 11, Fig. 4).
(C@N), 1578, 1542 (NH, C@C) cmÀ1 1H NMR:
.
d = 4.50 (s, 2H, COCH2), 6.48–8.19 (m, 4H, aromaticH),
9.61(s, 1H, NH). MS: m/z (%) 294, M+, (34.35) and 296,
M++2, (15.83). Anal. Found: C 49.20, H 2.70, N 9.43.
For C12H7ClN2O3S (294.72): C 48.91, H 2.39, N 9.51.
4.1.7.2. 2-Chloro-N-(7-oxo-7H-chromeno[6,5-d]thia-
zol-2-yl)propionamide 10b. Yield 85 %, crystallized from
ethanol, mp 237–239 ꢁC, IR: 3191 (NH), 3055 (CHa-
rom.), 2986, 2920, 2851 (CH3, CH), 1729, 1697 (2CO),
1
1619 (C@N), 1579, 1542 (NH, C@C) cmÀ1. H NMR:
Compounds, 3b, 6b, and 7b, that gave the highest pro-
tection at a dose of 100 mg/kg were studied at different
doses (50, 200 mg/kg) to calculate the PD50 which was
determined by log linear regression analysis from the
d = 1.69 (d, 3H, CH3), 4.86 (q, 1H, CH), 6.59 (d, 1H,
C-3H), 7.49 (d, 1H, C-7H), 7.98 (d, 1H, C-8H), 8.31
(d, 1H, C-4H), 12.94 (t, 1H, NH exchanged with

K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
5387
Table 11. Anticonvulsant activity of the tested compounds using PTZ
at a dose of 100 mg/kg and their log P values
vival time in min. The significance of differences
between groups was determined using one-way analysis
of variance (ANOVA) (Table 14).
Compound
Protection (%)
Log P
3b
3e
60
30
20
30
30
60
20
30
30
0
2.12 0.47
4.01 0.47
3.33 0.47
2.20 0.47
4.57 2.10
4.99 0.47
6.19 0.47
7.03 0.47
5.94 0.47
2.20 0.47
2.54 0.47
3.75 0.47
2.52 0.47
2.46 0.47
2.82 0.47
1.80 0.47
4.3. Determination of partition coefficient (log P)
3f
4b
ChemDraw Ultra 7.0 software was used for calculation
of log P of the tested compounds for antiepileptic
screening (Table 11).
5b
6b
6d
6e
4.4. Determination of ionization constant (pKa)
6g
7a
A spectrophotometric method was used for the determi-
nation of the pKa of biologically tested compounds 3b,
6b, and7b. The spectra of the ionized and unionized
forms were recorded in 0.1 M HCl and 0.1 M NaOH
with DMF. Wavelengths were chosen where the differ-
ence in absorbance between the ionized and unionized
forms was maximal (k = 336, 341, 273, and 324 nm,
respectively). The absorbance values at these wave-
lengths and at different pH (7.6, 7.4, 7, and 6.8) were
measured in a phosphate buffer (Na2HPO4/KH2PO4,
0.15 M) with DMF.42
7b
7e
80
60
0
8a
10a
0
11
Phenobarbital
40
90
dose response curves to compare with PD50 of phenbar-
bital (10, 20, and 30 mg/kg) (Table 12, Fig. 5).
4.2.2. Determination of LD50. Male albino mice weighing
25–30 g were divided into groups each of 8 animals. Pre-
liminary experiments were done to determine the mini-
mal dose that kills all animals (LD100), and the
maximal dose that fails to kill any animal. Several doses
at equal logarithmic intervals were chosen in between
these two doses, each dose was injected in a group of
eight animals, the number of dead animal in each group
after 24 h was recorded and the LD50 was calculated
according to Spearman Karber method.39 (Table 13) as
seen in the following formula
Table 13. Determination of the LD50 of compound 7b in PTZ test
Dose (mg/kg)
Log dose
No. of dead animals
400
800
2.60
2.90
3.20
3.50
0
2
3
8
1600
3200
Table 14. Anticonvulsant activity of the tested compounds using
strychnine at a dose of 100 mg/kg
M = Xk + 1/2 d À dr/N
M = Log LD50
Compound
Survival time (min) after strychnine
administration (M SD)
Xk = log dose causing 100% mortality.
d = logarithmic interval of doses.
r = Sum of the number of dead animals at each of the
individual dose levels.
Control
Phenytoin
3
1.10
13.8 4.50
3.92 1.20
5.02 1.15
5.33 0.516
5.40 0.548
4.85 1.19
5.18 2.21
7.55 1.14
4.50 0.495
8.36 2.44
5.00 0.935
3.14 1.09
6.26 1.23
4.40 0.946
5.44 2.31
3b
3e
3f
N = Number of animals at each of the dose level.
4.2.3. Strychnine seizure pattern test. Male albino mice,
weighing 25–30 g were divided into control group and
test groups (n = 6 per group). The control group re-
ceived 2% Tween 80 in water at a dose of 10 mL/kg po
while the test groups received the test compounds or
phenytoin at the dose of (100 mg/kg po) 45 min before
ip injection of strychnine (2 mg/kg). The average sur-
vival time (min) was recorded.40,41
4c
5b
5d
6b
6d
6g
7b
7e
8b
10a
11
Results of the experiments and observations are ex-
pressed as mean (M) standard deviation (SD) of sur-
Table 12. % Protection and PD50 of compounds 3b, 6b, 7b at dose 50, 100, and 200 mg and comparison with phenobarbital at dose 10, 20, and 30 mg
in PTZ test
50 mg
100 mg
200 mg
10 mg
20 mg
30 mg
PD50 (mg/kg)
3b
6b
20
20
30
—
60
60
80
—
70
60
90
—
—
—
—
40
—
—
—
70
—
—
—
90
125
138
95
7b
Phenobarbital
16

5388
K. M. Amin et al. / Bioorg. Med. Chem. 16 (2008) 5377–5388
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pKa = pH + log (AiÀA/AÀAu)
A = absorbance of the drug at a given pH
Ai = absorbance of the ionized drug
Au = absorbance of unionized drug
The mean value was then determined and standard devi-
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KON 941 spectrophotometer.
Acknowledgment
The authors are thankful to Dr. Hala Ahmed Fahmi,
Department of Pharmacology & Toxicology, Faculty
of Pharmacy, Cairo University, for carrying out the bio-
logical anticonvulsant screening part.
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