Design and synthesis of azolopyrimidoquinolines, pyrimidoquinazolines as anti-oxidant, anti-inflammatory and analgesic activities

Available online at www.sciencedirect.com

European Journal of Medicinal Chemistry 44 (2009) 609e624

http://www.elsevier.com/locate/ejmech

Original article

Design and synthesis of azolopyrimidoquinolines, pyrimidoquinazolines

as anti-oxidant, anti-inﬂammatory and analgesic activities

b

a

c

A.B.A. El-Gazzar ^a,, M.M. Youssef , A.M.S. Youssef , A.A. Abu-Hashem , F.A. Badria

*

^aPhotochemistry Department (Heterocyclic Unit), National Research Center, 12622 Dokki, Giza, Egypt

^bChemistry Department, Faculty of Science, Cairo University, Cairo, Egypt

^cDepartment of Pharmacognosy, Faculty of Pharmacy, Mansoura University, Mansoura, Egypt

Received 1 November 2007; received in revised form 17 March 2008; accepted 20 March 2008

Available online 8 April 2008

Abstract

The 5,10-dihydro-2-thioxo-pyrimido[4,5-b]quinolines (2aec) and its oxidized form 3 were prepared and used as key intermediates for the

synthesis of thiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]-quinolines (5aec), isoxazolo[5⁰⁰,4⁰⁰:4⁰,5⁰]thiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinolines (6aec),

4-chloro-2-methylthio-pyrimido[4,5-b]quinoline, its amino derivatives (19e21) and 10,11,12,13-tetrahydro-5H-quino[2⁰,3⁰:4,5]pyrimido[6,1-

b]quinazoline (22). The newly synthesized compounds were characterized by IR, NMR (¹H, ¹³C) and mass spectral studies. Representative

of the synthesized compounds was tested and evaluated for anti-oxidant, anti-inﬂammatory and analgesic activities. Compounds 2aec showed

the highest inhibitory anti-oxidant activity either using erythrocyte hemolysis or ABTS methods. Compounds 2a, 10b, 16, and 17a manifested

the best protective effect against DNA damage induced by bleomycin. Compounds 2c, 5a, 20a, 2a, and 2b exhibited a potent anti-inﬂammatory

activity using carrageenan-induced paw edema test in rats.

Keywords: Thiazolo[3⁰,2⁰:1,2]- and 2,3-dihydroisoxazolo[4⁰⁰,5⁰⁰:5⁰,4⁰]thiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinolones; Pyrimido[4,5-b]quinolone; 4-Amino deriva-

tives; Anti-oxidant; Anti-inﬂammatory; Analgesic activities

1. Introduction

in the present work the synthesis of some pyrimido[4,

5-b]quinoline, 2,10-diarylidene-7,8,9,10-tetrahydro-thiazolo

[3⁰,2⁰:1,2]pyrimido[4,5-b]quinoline-3,5-diones, 4-chloro-pyri-

mido[4,5-b]quinoline, and its 4-amino derivatives starting

from 6-amino-thiouracil and a,b-unsaturated ketones [18] in

order to investigate their anti-oxidant, analgesic and anti-

inﬂammatory activities.

Pyrimido[4,5-b]quinolones and their derivatives have

attracted interest over the years because of their varied biolog-

ical activities [1,2], recently found application in drug devel-

opment for the treatment of allergies [3], hypertension [4],

inﬂammation [5], central depressant agent [6], bacterial and

[7] HIV infections [8], antimalarial [9], anti-histaminic agents

[10], antitumor [11] and more recently for the treatment of

pain [12], antithrombotic activity [13] and as new inhibitions

of bacterial DNA gyrase B [14]. In view of the above men-

tioned ﬁndings and as continuation of our effort [15e17] to

identify new candidates that may be value in designing new,

potent, selective and less toxic antimicrobial agents, we report

2. Chemistry

Our group has actively been working on the development of

synthetic strategies for the preparation of 2-thioxo-pyrimido

[4,5-b]quinolin-4-ones (3) from a,b-unsaturated ketones 2

(Scheme 1). Thus, in a so-called cyclic strategy 5-(4-chloro-

phenyl)-9-(4-chlorophenylmethylene)-2-thioxo-6,7,8,9-tetrahy

dropyrimido[4,5-b]quinolin-4-one (3) was obtained by the re-

action of an a,b-unsaturated ketone (1) and 6-aminothiouracil

in reﬂuxing dimethylformamide for a long time. The structure

* Corresponding author. Tel./fax: þ20233700931.

E-mail address: profelgazzar@yahoo.com (A.B.A. El-Gazzar).

doi:10.1016/j.ejmech.2008.03.022

610

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

O

4 was prepared by the reaction of compound 3 with chloroacetic

acid in the presence of a mixture from acetic acid, acetic anhy-

dride and sodium acetate. Its IR spectrum displayed absorption

bands around 1696 and 1680 cm^ꢀ1for two carbonyl groups.

Compounds 5aec underwent cycloaddition with hydroxyl-

amine hydrochloride, by heating in boiling acetic acid in the

presence of sodium acetate, to give 7-arylidene-3,11-diaryl-

2,3,7,8,9,10-hexahydroisoxazolo[5⁰⁰,4⁰⁰:4⁰,5⁰]thiazolo[3⁰,2⁰:1,2]

pyrimido[4,5-b]quinolin-12-ones (6aec), with a new ring

Ar

O

CHAr

O

H

S

H

S

N

DMF

+

H

N

H₂N

N

H

N

H

Ar

1

H

Ar

O

Ar

O

H

N

2b

N

H

S

N

H

N

H

S

1

system. The H NMR spectrum of 6 showed a doublet signal

H

Ar

around 5.89e6.02 ppm which supported the oxazole proton,

furthermore the formation of 6 from 5 proceeded by ﬁrst

1,4-addition of hydroxylamine on the ethylenic double bond,

followed by loss of water [19].

H

Ar

2a-c

3

a, Ar = phenyl

b, Ar = 4-chlorophenyl

c, Ar = 4-methoxyphenyl

Ar = 4-Chlorophenyl

Alkylation of an ethanolic KOH solution of 3 with an alkyl

halide yielded the 2-alkylthio derivatives 7aee. Assignment of

structures 7 is based on the fact that each 7a,b gave the same 5-

aryl-2-hydrazino-9-arylmethylene-6,7,8,9-tetrahydropyrimido

[4,5-b]quinolin-4-one (8) with the evolution of methyl or ethyl

mercaptan on treatment with hydrazine hydrate (Scheme 3).

The ¹³C NMR (DMSO-d₆) spectrum for 7c showed the signals

at 16.41, 23.06, 24.21, 27.99, 28.25, 40.72 ppm for six sp³

carbons, and around 115.2e154.4 ppm for 16 sp²carbons

with four symmetric carbon atoms and at 167.57, 185.68 and

192.30 ppm assigned for three carbonyl groups.

On the other hand, trials to add hydrazine hydrate to 5 failed

and yielded instead the 2-hydrazino-8, which was reported

according to Shishoo [20], upon heating the 2-methylthio deriv-

ative with hydrazine hydrate. Assignments of structures 8 are

based on correct elemental analyses and IR and NMR

spectroscopies.

The reaction of 3 in an ethanolic potassium hydroxide so-

lution with a-haloketones, such as chloroacetone and/or

phenacylbromide, yielded 2-(S-acetone or S-phenacyl)-5-(4-

chlorophenyl)-9-(4-chlorophenylmethylene)-6,7,8,9-tetrahydr

opyrimido[4,5-b]quinolin-4-ones (7d,e) (Scheme 3). Assign-

ments of structures 7d,e are based on correct elemental ana-

lyses. The IR spectra are in agreement with the structure and

Scheme 1.

of 3 was established by analytical and spectral data [17], and

also the 5,10-dihydro derivatives 2aec were obtained after

reﬂuxing for 3e4 h.

In addition, we report here simple and convenient methods

for the syntheses of thiazolopyridopyrimidines, isoxazolothia-

zolopyridopyrimidines and pyridotriazolo-pyrimidines. Thus,

when a ternary mixture of 9-(4-chlorophenylmethylene)-5-(4-

chlorophenyl)-2-thioxo-2,3,6,7,8,9-hexahydropyrimido[4,5-b]-

quinolin-4-one (3), chloroacetic acid and a proper aldehyde was

heated under reﬂux in a mixture of acetic acid, acetic anhydride

and anhydrous sodium acetate, 6-aryl-2,10-diarylmethylene-

7,8,9,10-tetrahydrothiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quino-

line-3,5-diones (5aec) were obtained in high yields (Scheme 2).

The structure assignments were based on an independent

preparation of 5a by condensation of 6-aryl-10-arylmethy-

lene-7,8,9,10-tetrahydrothiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]-

quinoline-3,5-dione 4 with aldehyde (Scheme 2) and the correct

values in elemental analysis and compatible spectral data.

The ¹³C NMR spectrum for compound 5c showed the signals

at 24.90, 30.72, 30.95 ppm for three sp³carbons, 115.6e

155.6 ppm for 22 sp²carbons with four symmetric carbon

atoms, and 166.3 and 168.6 (2CO) ppm. Moreover, compound

revealed the presence of

a

free keto-group around

1720 cm^ꢀ1. The latter compounds 7d,e were cyclized under

reﬂux in a mixture of glacial acetic acid and sulfuric acid to

give 3-(methyl or phenyl)-6-(4-chlorophenyl)-10-(4-chloro-

phenylmethylene)-7,8,9,10-tetrahydrothiazolo[3⁰,2⁰:1,2]pyrim

ido[4,5-b]quinoline-3,5-diones (9a,b). Structures 9a,b were

preferred on the basis of ¹H and ¹³C NMR spectral data.

Also the IR spectra are in agreement with the structure and re-

vealed the absence of the NH group at position 3. The 2-al-

kylthio derivatives 7a,b underwent further alkylation at the

N-3 nitrogen atom on treatment with alkyl iodides, in aqueous

ethanolic sodium ethoxide solution, to afford 2-alkylthio-3-

alkyl-5-(4-chlorophenyl)-9-(4-chlorophenylmethylene)-6,7,8,9-

tetra-hydropyrimido[4,5-b]quinolin-4-ones (10aed). Also,

oxidation of 7a with hydrogen peroxide in acetic acid yielded

the 2-methylsulfone (11) (Scheme 3). Structures 11 were pre-

ferred from analytical data and spectroscopic analysis and also

on the shifting of the 2-methylthio group at position 2 from

2.84 to 3.30 ppm.

Ar

N

O

Ar'-CHO

N

3 + ClCH₂COOH

AcOH/Ac₂O

AcOH/Ac O

NaOAc²

N

S

CHAr'

5a-d

NaOAc

Ar

H

NH₂OH

aldehyde

NaOAc

Ar

N

Ar

O

N

O

H

N

S

N

S

Ar'

Ar

H

Ar

4

6a-c

Ar = 4-chlorophenyl

b, Ar' = 4-methoxyphenyl

d, Ar' = 4- N,N-dimethylaminophenyl

;

a, Ar' = phenyl

c, Ar' = 2-thienyl

Scheme 2.

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

611

Ar

O

Ar

N

R

O

N

R¹

S

7d,e

7a,b

N

S

N

sulphuric acid

AcOH

R

alkyl-iodide

N

H

Ar

R = CH₃, C₆H₅

10a-d

H

Ar

O

9a,b

H₂O₂

NH

alkyl-iodide

3a

R

ethanol/KOH

N

S

Ar

O

H

N

Ar

H

7a,b

Ar

NH₂NH₂.H₂O

O

CH₃

7a-e

S

N

O

H

N

Ar

H

11

10a, R = R' = CH₃

10b, R = CH₃, R' = C₂H₅

7a, R = CH₃

7b, R = C₂H₅

NH₂

N

H

7c, R = CH(COCH₃)₂

7d, R = CH₂COCH₃

7e, R = CH₂COC₆H₅

10c, R = C₂H₅, R' = CH₃

10d, R = R' = C₂H₅

Ar

8

H

Ar = 4-Chlorophenyl

Scheme 3.

On the other hand, compound 7c, as typical 1,3-diketones,

reacted with hydrazine hydrate and thiourea to afford the cor-

responding 2-(3,5-dimethyl-1H-pyrazol-4-yl-thio) and 2-(4,6-

dimethyl-2-thioxo-pyrimidin-5-yl-thio) derivatives 12 and 13,

respectively (Scheme 4). Heating 7c under reﬂux on acetic an-

hydride/pyridine mixture led to cyclization and formation of

2-acetyl-6-(4-chlorophenyl)-10-(4-chlorophenylmethylene)-3-

methyl-7,8,9,10-tetrahydro-thiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]

quionolin-5-one (14) in good yield. Assignments of structures

14 were based on spectral data besides our previous report on

a related compound [19]. Thus the N-3 nitrogen atom and not

the N-1 nitrogen atom was involved in the cyclization. The ¹³C

NMR spectrum of 14 showed the signals at 12.70, 24.81 (2C,

2CH₃) ppm, 25.72, 29.91, 30.13 (3C, 3CH₂) ppm, 18 sharp

lines around 104.2e155.1 ppm supported the presence of 22

sp²carbon atoms (included four symmetric carbons of the

Ar

O

Ar

O

N

H₃C

S

H₃C

NH

N

S

N

NH

H

N

S

CH₃

thio-

urea

CH₃

Ar

H

N₂H₄.H₂O

13

12

Ar

H

Ar

O

Ar

O

N

CH₃

H

Ac₂O

7c

CH₃

COCH₃

N

pyridine

N

Ar'

N

S

N

S

Ar'-CHO

O

H

Ar

H

Ar

H

14

17a, b

a, Ar' = phenyl

NH₂OH

Thiosemicarbazide

b, Ar' = 4-methoxyphenyl

Ar

O

Ar

O

CH₃

N

S

NNHCSNH₂

CH₃

N

S

C

N

C

CH₃

Ar

H

NOH

Ar

H

15

16

Ar = 4-chlorophenyl

Scheme 4.

612

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

p-substituted phenyl) and two signals at 165.3, 193.1 due to

the two carbonyl groups.

[20] and its activity towards nucleophiles such as primary

aromatic amines, secondary amines and hydrazine hydrates

was investigated. Thus heating compound 18 under reﬂux

with aryl-amine, namely aniline, 4-chloroaniline and p-anisi-

dine in acetic acid, produced the 4-arylamino-2-methylthio-5-

(4-chlorophenyl)-9-(4-chlorophenylmethylene)-6,7,8,9-tetrahy

dropyrimido[4,5-b]quinolines (19aec) (Scheme 5).

In support of structure 14, characteristic reactions for the

2-acetyl group were observed. Thus, its reaction with each

of hydroxylamine hydrochloride and thiosemicarbazide

gave the corresponding oxime and thiosemicarbazone

derivatives 15 and 16, respectively (Scheme 4). Compounds

15 and 16 gave correct values in elemental analyses and

The IR spectrum of 18 revealed the absence of any absorp-

tion bands in the NH region. The IR spectra of 19 displayed

an absorption band at 3395e3420 cm^ꢀ1(NH). Moreover, the

reaction of 4-chloropyridopyrimidine (18) with anthranilic

acid afforded 4-(o-carboxyphenyl)-amino-pyrimidoquinoline

(21) (Scheme 5). The IR spectra of 21 displayed an absorption

band at 3510 cm^ꢀ1(OH), 3368 cm^ꢀ1(NH), and 1716 cm^ꢀ1for

carbonyl group. The latter compound 21 underwent cyclization

when boiled with glacial acetic acid in the presence of catalytic

amount of sulfuric acid to give 14-(4-chlorophenyl)-10-

(4-chlorophenylmethylene)-7-methylthio-10,11,12,13-tetrahy-

dro-5H-quino[2⁰,3⁰:4,5]pyrimido[6,1-b]quinazolin-5-one (22).

1

compatible data in IR and H NMR spectra. The mass spec-

trum of 14 showed a molecular ion peak at m/z 545 for [M^þ]

(100%). Moreover, compound 14 yielded the 6-aryl-10-aryl-

methylene-2-cinnamoyl-3-methyl-7,8,9,10-tetrahydrothiazo-

lo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinolin-5-one derivatives 17a,b

on heating with the proper aldehyde in the presence of

a catalytic amount of piperidine (Scheme 5). The IR spectra

of 17 displayed two carbonyl absorption bands around 1700e

1

1685 cm^ꢀ1. Also the H NMR of 17 revealed the presence

two doublet signals supporting the AB system hydrogen

atoms in the trans-form for the ethylenic protons around

5.28e5.53 with J coupling around 10.9, 11.5 Hz.

1

Structure 22 was preferred on the basis of H NMR and ¹³C

Finally, it is well known in pyrimidine chemistry that

position 4 in pyrimidines and fused pyrimidines shows distinct

activities towards nucleophiles. Therefore, 4-chloro-2-methyl-

thio-pyrimido[4,5-b]quinoline (18) was prepared from the

reaction of 7a with phosphorus oxychloride in dry dioxane

NMR spectral data. Similarly, the 2,4-dihydrazinopyridopyri-

midine derivative 23 was synthesized from 18 and hydrazine

hydrate by heating in dry dioxane. Beside the correct values

in elemental analyses spectral data of 23 are in agreement

with the assigned structure.

7a

X

Ar

N

HN

N

POCl₃

N

Ar

N

Cl

Aryl-amine

19a, X = H

19b, X = Cl

19c, X = OMe

SCH₃

N

19a-c

Ar

H

N

SCH₃

X

Ar

H

18

Ar

N

NH₂NH₂

N

2^ndamine

Ar

NHNH₂

20a, X = NH

20b, X = O

SCH₃

N

Ar

H

20a, b

N

NHNH₂

HOOC

anthranilic

acid

23

Ar

H

HN

Ar

N

Ar

N

sulphuric acid

N

O

SCH₃

N

SCH₃

N

Ar

H

Ar

H

21

22

Ar = 4-chlorophenyl

Scheme 5.

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

613

Table 2

Anti-oxidant assays by ABTS method [Abs (control) ꢀ Abs (test)/Abs

3. Results and discussion

(control) ꢁ 100]

Compounds

3.1. Anti-oxidant activity screening

Absorbance

of samples

Inhibition

(%)

A variety of quinoline compound derivatives were tested for

anti-oxidant activity as reﬂected in the ability to inhibit lipid per-

oxidation in rat brain and kidney homogenates and rate erythro-

cyte hemolysis. The pro-oxidant activities of the aforementioned

compounds were assessed by their effects on bleomycin-induced

DNA damage. The pyrimido[4,5-b]quinoline derivatives mani-

fested potent anti-oxidative activity in the lipid peroxidation

assay but showed no inhibitory activity in the hemolysis assay.

All compounds have been tested to bleomycin-dependant

DNA damage. The results indicated that they may have some

protective activity to DNA by certain mechanism. A series of

compounds (2a, 2c, 2b) exhibited a high anti-oxidant activity.

On the other hand, compounds 2a, 10b, 16, 17a protect the

DNA from the induced damage by bleomycin (Tables 1e4).

Control of ABTS

Ascorbic acid

2a

0.54

0.06

0.10

0.22

0.18

0.30

0.32

3.36

0.40

0.34

0.36

0.41

0.38

0.36

0.34

0.31

0.35

0.36

0.37

0.34

0.37

0.35

0.32

0.28

0.20

0.33

0.42

0.34

0

88.9

81.5

59.3

66.7

44.4

40.7

33.3

25.9

37.0

33.3

24.1

29.6

33.3

37.0

42.6

35.2

33.3

31.5

37.0

31.5

35.2

40.7

48.1

63.0

38.9

22.2

37.0

2b

2c

3

5a

5b

5c

5d

6a

6b

6c

7a

7b

7c

7d

7e

10a

10b

10c

10d

14

3.2. Anti-inﬂammatory activity

Anti-inﬂammatory activity was evaluated by carrageenan-

induced paw edema test in rats. The anti-inﬂammatory activity

data (Table 5) indicated that all the test compounds protected

rats from carrageenan-induced inﬂammation and some of the

tested compounds are more potent than our earlier reported

15

16

17a

17b

18

Table 1

Anti-oxidant assays by erythrocyte hemolysis (A/B ꢁ 100)

compounds [21]. Compounds 2aec, 5a, 6a, 7a, 20a showed sim-

ilar and more anti-inﬂammatory activity than diclofenac sodium.

Compounds

Absorbance

of samples (A)

Hemolysis

(%)

Complete hemolysis

0.660

with distilled H₂O (B)

Ascorbic acid

2a

3.3. Analgesic activity

0.026

0.236

0.084

0.054

0.055

0.038

0.030

0.042

0.048

0.062

0.048

0.031

0.028

0.038

0.052

0.055

0.043

0.027

0.026

0.035

0.203

0.040

0.048

0.041

0.030

0.039

0.029

3.93

35.75

12.72

8.18

8.33

5.75

4.54

6.36

7.27

9.39

7.27

4.64

4.24

5.60

7.87

8.33

6.51

4.09

3.93

5.30

30.75

6.06

7.22

6.21

4.54

5.90

4.39

Test for analgesic activity was performed by Turner [28] and

Collier [29] technique using Swiss albino mice. The results of

analgesic activity indicated that all test compounds exhibited

signiﬁcant activity. Compounds 2aec, 5a, 6a, 20a,b have nearly

the same activity of reference drug, and the remaining tested

compounds have good central analgesic activity (Table 6).

Also dihydropyrimido[4,5-b]quinolines (2aec) and thiazo-

lo[3⁰,2⁰:1,2]pyrimido[4,5-b]-quinoline-3,5-dione (5a) have good

activity higher than the reference drug in peripheral analgesic ac-

tivity testing. Isoxazolothiazolo (6a), 2-alkylthio- (7a,c), 4-piper-

azino (20a) and 4-morpholino-pyrimido[4,5-b]quinolines (20a,b)

have the same potency as reference drug. Furthermore, the

2b

2c

3

5a

5b

5c

5d

6a

6b

6c

7a

7b

7c

7d

7e

10a

10b

10c

10d

14

Table 3

Assay for bleomycin-dependent DNA damage (DNA)

Compounds

Absorbance

of samples

15

16

Ascorbic acid

2a

0.020

0.025

0.021

0.065

0.035

17a

17b

18

10b

16

17a

614

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

Table 4

Comparison between dihydrocompounds 2aec in anti-oxidant assays by

ABTS method

5. Experimental

5.1. Chemistry

2a

H

2b

Cl

2c

OCH₃

ABTS (%)

66.70

All melting points were measured using an Electrothermal

IA 9100 apparatus (Shimadzu, Japan). Microanalytical data

were recorded by Vario, Elementar apparatus (Shimadzu)

(Table 8). The IR spectra (KBr) were recorded on a Perkine

Elmer 1650 spectrometer (USA). ¹H NMR spectra were

determined on a JEOL EX-270 run for H NMR at 270 MHz

and run for C NMR at 67.5 MHz: or on JEOL ECA-500 run

for H NMR at 500 MHz and run for C NMR at 125 MHz,

and chemical shifts were expressed in parts per million relative

to SiMe₄as internal standards. Mass spectra were recorded on

70 eV EI Ms-QP 1000 EX (Shimadzu, Japan). The starting

materials are prepared according to El-Gazzar [18].

ABTS (%)

81.50

H (%)

35.75

ABTS (%)

59.30

H (%)

12.72

H (%)

8.18

remaining compounds have moderate activity in peripheral anal-

gesic activity testing (Table 7).

4. Conclusions

The prepared new ring systems seem to be interesting for

biological activity studies. Furthermore, the present investiga-

tion offers rapid and effective new procedures for the synthesis

of the polycondensed new heterocyclic ring systems.

Compounds 2aec showed the highest inhibitory anti-oxidant ac-

tivity either using erythrocyte hemolysis or ABTS methods.

These quinolones are either phenyl, or 4-chlorophenyl or 4-

methoxyphenyl. This means the presence of the phenyl group

will potentiate the activity which may increase by introducing

an electron-donating group e.g. OCH₃, or electron-withdrawing

groupe.g. Clinp-position. Compounds 2a, 10b, 16and17aman-

ifestedthebestprotectiveeffectagainstDNAdamageinducedby

bleomycin. The phenyl derivative of 2 proved to be more active

than that of methoxy or chloro derivatives. Compounds 2c, 5a,

20a, 2a and 2b exhibited a potent anti-inﬂammatory activity us-

ing carrageenan-induced paw edema test in rats. The p-methoxy

derivative of 2 retained the best activity in all tested assays. This

may suggest that the mechanism of anti-inﬂammatory and anal-

gesic activities of compounds 2aec, especially 2c, may be due to

the potent anti-oxidant activity to maintain the integrity of both

the cell membrane and DNA of the cells.

5.1.1. Synthesis of 5-aryl-9-(arylmethylene)-5,10-dihydro-2-

thioxo-6,7,8,9-tetrahydropyrimido[4,5-b]quinolin-4-one

(2aec): general procedure

A mixture of 1aec (0.01 mol) and 6-aminothiouracil

(0.01 mol) was reﬂuxed in dimethylformamide (50 mL) for

3e4 h. The reaction mixture was cooled, the deposited precip-

itate was ﬁltered off, washed with ethanol and dried, and

crystallized from dimethylformamide to obtain 2aec, as

yellow powder products.

5.1.1.1. Synthesis of 5,10-dihydro-5-phenyl-9-(phenylmethy-

lene)-2-thioxo-6,7,8,9-tetrahydropyrimido[4,5-b]quinolin-

4-one (2a). It was obtained from 1a. IR (KBr, cm^ꢀ1): 3385

(br, NH’s), 3056 (CH aryl), 2924 (CH alkyl), 1686 (CO),

1

1635 (C]N); H NMR (DMSO-d₆, d, ppm): 1.65e1.67 (m,

2H, CH₂), 2.29 (t, 2H, J ¼ 5.83 Hz, CH₂), 2.80 (t, 2H, J ¼

5.85 Hz, CH₂), 5.43 (s, 1H, C5eH), 7.00e7.13 (m, 2H, phe-

nyl), 7.16e7.19 (m, 5H, phenyl), 7.25e7.28 (m, 3H, phenyl),

8.21 (s, 1H, CH), 9.50, 10.40, 12.00 (3 br s, 3NH, D₂O

exchangeable); its MS, [M^þ], m/z 399 (100%).

Table 5

Percent inﬂammatory activity of the tested compounds (carrageenan-induced

paw edema in rats)

Compounds

Percent protection

1 h

5.1.1.2. Synthesis of 5-(4-chlorophenyl)-9-(4-chlorophenyl-

methylene)-5,10-dihydro-2-thioxo-6,7,8,9-tetrahydropyr-

imido[4,5-b]quinolin-4-one (2b). It was obtained from 1b. IR

(KBr, cm^ꢀ1): 3390 (br, NH’s), 3036 (CH aryl), 2941 (CH

alkyl), 1690 (CO), 1615 (C]N); ¹H NMR (DMSO-d₆, d,

ppm); 1.63e1.66 (m, 2H, CH₂), 2.28 (t, 2H, J ¼ 5.78 Hz,

CH₂), 2.78 (t, 2H, J ¼ 5.83 Hz, CH₂), 5.41 (s, 1H, C5eH),

7.12 (d, 2H, J ¼ 8.35 Hz, phenyl), 7.17 (d, 2H, J ¼ 8.37 Hz,

phenyl), 7.22e7.28 (m, 4H, phenyl), 8.21 (s, 1H, CH),

10.20, 11.00, 12.30 (3 br s, 3NH, D₂O exchangeable); its

MS, [M^þ], m/z 467 (100%), [M^þþ 2], m/z 469 (52%),

[M^þþ 4], m/z 471 (13%).

2 h

3 h

2a

2b

49.0 ꢂ 1.27*

56.3 ꢂ 1.37**

59.5 ꢂ 1.06**

59.8 ꢂ 1.41**

49.6 ꢂ 2.41*

50.1 ꢂ 1.51**

46.3 ꢂ 1.53*

46.7 ꢂ 2.28*

41.2 ꢂ 1.38*

42.3 ꢂ 1.63*

44.1 ꢂ 1.83*

39.8 ꢂ 1.42*

43.1 ꢂ 162*

39.2 ꢂ 1.39*

34.0 ꢂ 1.36*

6.1 ꢂ 0.27

53.2 ꢂ 1.32**

62.0 ꢂ 1.76**

62.4 ꢂ 2.03**

59.3 ꢂ 1.32*

57.1 ꢂ 1.63*

56.2 ꢂ 1.64**

48.6 ꢂ 139*

44.2 ꢂ 1.83*

42.5 ꢂ 1.46*

45.8 ꢂ 1.47*

49.0 ꢂ 1.14

41.0 ꢂ 1.31*

40.2 ꢂ 1.05*

46.5 ꢂ 1.26*

42.3 ꢂ 1.26*

34.8 ꢂ 1.33*

32.4 ꢂ 1.21*

38.6 ꢂ 1.39*

39.2 ꢂ 1.04*

31.1 ꢂ 1.33*

32.9 ꢂ 1.26*

30.5 ꢂ 1.83*

25.3 ꢂ 1.32*

42.0 ꢂ 1.92*

37.3 ꢂ 1.21*

36.7 ꢂ 1.54*

3.2 ꢂ 0.93

2c

5a

6a

7a

7c

14

18

19a

19b

19c

42.3 ꢂ 1.42*

52.0 ꢂ 1.95**

47.1 ꢂ 1.45*

39.6 ꢂ 1.46*

5.7 ꢂ 0.32

20a

20b

5.1.1.3. Synthesis of 5-(4-methoxyphenyl)-9-(4-methoxyphenyl-

methylene)-5,10-dihydro-2-thioxo-6,7,8,9-tetrahydropyr-

imido[4,5-b]quinolin-4-one (2c). It was obtained from 1c. IR

(KBr, cm^ꢀ1): 3410 (br, NH’s), 3047 (CH aryl), 2916 (CH

alkyl), 1686 (CO), 1630 (C]N); ¹H NMR (DMSO-d₆, d,

ppm): 1.61e1.65 (m, 2H, CH₂), 2.24 (t, 2H, J ¼ 5.84 Hz, CH₂),

23

Control

Diclofenac sodium

52.4 ꢂ 0.92*

60.3 ꢂ 1.52**

42.0 ꢂ 1.36*

Each value represents the mean ꢂ SE (n ¼ 6). Signiﬁcance levels *p < 0.5,

**p < 0.001 as compared with respective control. Dose is 20 mg/kg for the

selected tested compound.

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

615

Table 6

Central analgesic activity (hot-plate test)

Compounds

Reaction time (min)

0 min

30 min

60 min

90 min

Control

2a

8.23 ꢂ 0.33

9.40 ꢂ 0.35

7.40 ꢂ 0.38

8.16 ꢂ 0.57

8.90 ꢂ 0.65

6.24 ꢂ 0.57

7.40 ꢂ 0.36

8.26 ꢂ 0.40

6.64 ꢂ 0.20

8.26 ꢂ 0.40

7.26 ꢂ 0.40

9.38 ꢂ 0.30

8.42 ꢂ 0.61

6.08 ꢂ 0.90

7.08 ꢂ 0.13

8.09 ꢂ 0.34

6.49 ꢂ 0.40

8.10 ꢂ 0.36^b

9.42 ꢂ 0.45^a

9.84 ꢂ 0.28^a

9.16 ꢂ 0.57^a

7.08 ꢂ 0.78^a

8.42 ꢂ 0.45^a

8.68 ꢂ 0.48

7.54 ꢂ 0.26^b

9.68 ꢂ 0.48

8.50 ꢂ 0.48

10.14 ꢂ 0.26^b

8.62 ꢂ 0.34

7.93 ꢂ 0.74

8.65 ꢂ 0.87

8.82 ꢂ 0.35^a

10.03 ꢂ 0.12^a

8.60 ꢂ 0.41^b

10.41 ꢂ 0.29^a

11.62 ꢂ 0.57^a

10.78 ꢂ 0.45^a

9.52 ꢂ 0.82^a

10.41 ꢂ 0.29^a

9.80 ꢂ 0.52

9.51 ꢂ 0.40^b

11.67 ꢂ 0.59^a,b

11.88 ꢂ 0.47^a

11.08 ꢂ 0.24^b

12.68 ꢂ 0.61^a

10.67 ꢂ 0.59^a,b

10.40 ꢂ 0.18^b

12.00 ꢂ 0.36^a

10.40 ꢂ 0.18^b

10.47 ꢂ 0.18^b

7.62 ꢂ 0.60^b

2b

2c

5a

6a

7a

7c

14

18

8.61 ꢂ 0.60^a

9.80 ꢂ 0.52

19a

19b

9.86 ꢂ 0.52

10.74 ꢂ 0.28^b

9.06 ꢂ 0.56^b

10.15 ꢂ 1.20

10.98 ꢂ 0.91^a

9.86 ꢂ 0.58^a

11.39 ꢂ 0.53^a

19c

20a

9.22 ꢂ 0.47^b

11.50 ꢂ 0.40^a

11.12 ꢂ 0.75^a,b

10.46 ꢂ 0.48^b

13.15 ꢂ 0.38^a

20b

23

Diclofenac sodium

Values represent the mean ꢂ SE of six animals for each group.

a

p < 0.05: statistically signiﬁcant from control (Dunnett’s test).

p < 0.05: statistically signiﬁcant from ASA (Dunnett’s test).

b

2.73 (t, 2H, J ¼ 5.79 Hz, CH₂), 3.87, 3.91 (2s, 6H, 2CH₃), 5.45

(s, 1H, C5eH), 7.14 (d, 2H, J ¼ 8.42 Hz, phenyl), 7.20 (d, 2H,

J ¼ 8.43 Hz, phenyl), 7.26e7.29 (m, 4H, phenyl), 8.30 (s, 1H,

CH), 9.20, 10.20, 11.50 (3 br s, 3NH, D₂O exchangeable); its

MS, [M^þ], m/z 459 (100%).

20e30 h. The reaction mixture was cooled, the deposited pre-

cipitate was ﬁltered off, washed with ethanol and dried, and

crystallized from dimethylformamide. IR (KBr, cm^ꢀ1): 3380

(br, NH’s), 3025 (CH aryl), 2921 (CH alkyl), 1688 (CO),

1

1631 (C]N); H NMR (DMSO-d₆, d, ppm): 1.64e1.67 (m,

2H, CH₂), 2.30 (t, 2H, J ¼ 6.00 Hz, CH₂), 2.75e2.97 (t, 2H,

J ¼ 5.93 Hz, CH₂), 7.11 (d, 2H, J ¼ 8.40 Hz, phenyl), 7.15

(d, 2H, J ¼ 8.39 Hz, phenyl), 7.17e7.19 (m, 4H, phenyl),

8.19 (s, 1H, CH), 10.30, 11.50 (2 br s, 2NH); ¹³C NMR

(DMSO-d₆, d, ppm): 22.27, 26.73, 27.18 (3C, 3CH₂), 108.3,

127.9, 128.2, 128.5, 128.6, 129.4, 130.1, 131.1, 132.3,

5.1.2. Synthesis of 5-(4-chlorophenyl)-9-

(4-chlorophenylmethylene)-2-thioxo-6,7,8,9-

tetrahydropyrimido[4,5-b]quinolin-4-one (3)

A mixture of 1b (0.01 mol) and 6-aminothiouracil (0.01

mol) was reﬂuxed in dimethylformamide (50 mL) for

Table 7

Percent analgesic activity (peripheral, writhing test)

Compounds

Percent protection

30 min

1 h

2 h

3 h

2a

2b

61.2 ꢂ 1.15*

60.4 ꢂ 1.51**

69.0 ꢂ 1.05**

73.2 ꢂ 1.05**

46.7 ꢂ 1.51*

50.4 ꢂ 1.34*

46.0 ꢂ 1.93*

43.5 ꢂ 1.92*

41.4 ꢂ 1.16*

43.4 ꢂ 1.21*

43.2 ꢂ 1.32*

41.5 ꢂ 1.43*

49.5 ꢂ 1.05*

45.2 ꢂ 1.46*

41.0 ꢂ 1.63*

02.0 ꢂ 0.35

65.1 ꢂ 1.31**

69 ꢂ 1.56**

74 ꢂ 1.93**

76 ꢂ 1.39**

53 ꢂ 1.49**

52 ꢂ 1.02*

54 ꢂ 1.41*

49 ꢂ 1.37*

47 ꢂ 1.32*

48 ꢂ 1.42*

45 ꢂ 1.03*

52 ꢂ 1.26*

50 ꢂ 1.63*

47 ꢂ 1.52*

45 ꢂ 1.32*

06.0 ꢂ 0.50

55.2 ꢂ 1.16*

69.2 ꢂ 1.03**

74.3 ꢂ 1.84**

75.4 ꢂ 1.51**

77.2 ꢂ 1.31

59.3 ꢂ 1.69

55.5 ꢂ 1.37

56.6 ꢂ 1.39

53.1 ꢂ 1.16

48.8 ꢂ 1.39

51.3 ꢂ 1.60

46.1 ꢂ 1.39

49.3 ꢂ 1.92

49.4 ꢂ 1.37

45.2 ꢂ 1.47

49.7 ꢂ 1.92

04.0 ꢂ 0.59

62 ꢂ 1.49*

50.1 ꢂ 1.73*

47.3 ꢂ 172*

58.2 ꢂ 1.17*

60.3 ꢂ 1.39**

36.5 ꢂ 1.09*

37.6 ꢂ 1.49*

37.3 ꢂ 1.23*

38.4 ꢂ 1.81*

36.2 ꢂ 1.74*

39.6 ꢂ 1.83*

34.7 ꢂ 1.15*

37.3 ꢂ 1.39*

30.5 ꢂ 1.61*

27.8 ꢂ 1.43*

33.1 ꢂ 1.32*

04.0 ꢂ 0.90

39 ꢂ 1.13*

2c

5a

6a

7a

7c

14

18

19a

19b

19c

20a

20b

23

Control

Diclofenac sodium

46.0 ꢂ 095*

Each value represents the mean ꢂ SE (n ¼ 6). Signiﬁcance levels *p < 0.5, **p < 0.001 as compared with respective control. Dose is 20 mg/kg for the selected

tested compound.

616

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

Table 8

Physical and chemical properties of synthesized compounds

Compounds

Yield (%)

M.p. (^ꢃC)

Mol. formula (M_r)

Found (calculated) (%)

C

H

N

2a

66

64

67

85

73

85

80

81

82

80

74

85

88

90

80

82

80

75

77

78

76

71

58

60

80

70

80

61

65

80

74

76

68

63

77

75

90

330e332

215e217

228e230

295e297

302e304

354e356

330e332

316e218

195e197

238e240

188e190

290e292

327e330

310e312

239e241

205e207

228e230

278e280

290e292

272e274

218e220

245e247

146e148

168e170

243e246

270e273

261e264

221e223

269e271

181e183

289e291

256e259

360e362

245e247

282e285

257e260

227e230

241e243

223e225

184e186

298e300

C₂₄H₂₁N₃OS (399.5)

C₂₄H₁₉Cl₂N₃OS (468.4)

C₂₆H₂₅N₃O₃S (459.5)

72.15 (72.17)

61.54 (61.56)

67.95 (67.94)

61.81 (61.78)

61.66 (61.64)

66.66 (66.70)

65.38 (65.40)

62.00 (62.04)

65.93 (65.95)

65.02 (65.00)

63.85 (63.88)

60.48 (60.51)

62.50 (62.47)

63.15 (63.17)

61.70 (61.68)

62.07 (62.05)

65.76 (65.73)

62.08 (62.11)

64.26 (64.27)

67.84 (67.82)

63.15 (63.12)

63.78 (63.76)

63.78 (63.80)

64.36 (64.40)

58.60 (58.61)

62.14 (62.09)

59.60 (59.58)

63.74 (63.75)

62.03 (62.02)

58.16 (58.18)

68.14 (68.12)

66.86 (66.84)

60.19 (60.17)

67.03 (67.01)

63.11 (63.09)

65.64 (65.62)

63.97 (64.01)

63.85 (63.82)

64.10 (64.09)

66.09 (66.11)

60.25 (60.23)

5.30 (5.27)

4.09 (4.11)

5.48 (5.46)

3.67 (3.65)

3.38 (3.37)

3.56 (3.59)

3.71 (3.68)

3.19 (3.22)

4.11 (4.13)

3.64 (3.67)

3.78 (3.80)

3.27 (3.29)

3.98 (3.94)

4.28 (4.30)

4.10 (4.08)

4.05 (4.03)

3.96 (3.98)

4.12 (4.09)

3.79 (3.80)

3.74 (3.71)

4.28 (4.31)

4.56 (4.54)

4.56 (4.60)

4.82 (4.85)

3.74 (3.72)

4.13 (4.15)

3.83 (3.85)

3.87 (3.90)

3.95 (3.97)

3.90 (3.87)

3.97 (3.95)

4.10 (4.12)

3.63 (3.65)

4.35 (4.29)

3.93 (3.90)

4.47 (4.45)

4.26 (4.23)

4.07 (4.05)

4.03 (4.01)

3.81 (3.79)

4.43 (4.45)

10.52 (10.49)

8.97 (8.85)

9.14 (9.16)

9.01 (9.06)

8.29 (8.30)

7.09 (7.12)

6.73 (6.75)

6.99 (7.01)

8.79 (8.83)

9.19 (9.20)

8.76 (8.75)

9.10 (9.12)

8.75 (8.79)

8.50 (8.46)

7.44 (7.41)

8.04 (8.01)

7.19 (7.20)

15.08 (15.06)

8.33 (8.35)

7.42 (7.45)

8.50 (8.48)

8.26 (8.30)

8.26 (8.29)

8.04 (8.02)

8.20 (8.17)

12.49 (12.47)

11.58 (11.61)

7.69 (7.71)

9.98 (10.01)

13.57 (13.49)

6.62 (6.58)

7.22 (7.19)

8.42 (8.43)

10.09 (10.07)

9.50 (9.47)

9.57 (9.59)

12.86 (12.84)

10.27 (10.30)

9.35 (9.37)

9.63 (9.65)

20.50 (20.52)

2b

2c

3

C₂₄H₁₇Cl₂N₃OS (466.3)

C₂₆H₁₇Cl₂N₃O₂S (506.4)

C₃₃H₂₁Cl₂N₃O₂S (594.5)

C₃₄H₂₃Cl₂N₃O₃S (624.5)

C₃₁H₁₉Cl₂N₃O₂S₂(600.5)

C₃₅H₂₆Cl₂N₄O₂S (637.5)

C₃₃H₂₂Cl₂N₄O₂S (609.5)

C₃₄H₂₄Cl₂N₄O₃S (639.5)

C₃₁H₂₀Cl₂N₄O₂S₂(615.5)

C₂₅H₁₉Cl₂N₃OS (480.4)

C₂₆H₂₁Cl₂N₃OS (494.4)

C₂₉H₂₃Cl₂N₃O₃S (564.5)

C₂₇H₂₁Cl₂N₃O₂S (522.4)

C₃₂H₂₃Cl₂N₃O₂S (584.5)

C₂₄H₁₉Cl₂N₅O (464.4)

C₂₇H₁₉Cl₂N₃OS (504.4)

C₃₂H₂₁Cl₂N₃OS (566.5)

C₂₆H₂₁Cl₂N₃OS (494.4)

C₂₇H₂₃Cl₂N₃OS (508.4)

C₂₈H₂₅Cl₂N₃OS (522.5)

C₂₅H₁₉Cl₂N₃O₃S (512.4)

C₂₉H₂₃Cl₂N₅OS (560.5)

C₃₀H₂₃Cl₂N₅OS₂(604.5)

C₂₉H₂₁Cl₂N₃O₂S (546.4)

C₂₉H₂₂Cl₂N₄O₂S (561.5)

C₃₀H₂₄Cl₂N₆OS₂(619.6)

C₃₆H₂₅Cl₂N₃O₂S (634.5)

C₃₇H₂₇Cl₂N₃O₃S (664.6)

C₂₅H₁₈Cl₃N₃S (498.8)

C₃₁H₂₄Cl₂N₄S (555.5)

C₃₁H₂₃Cl₃N₄S (589.9)

C₃₂H₂₆Cl₂N₄OS (585.5)

C₂₉H₂₃Cl₂N₅S (544.5)

C₂₉H₂₂Cl₂N₄OS (545.5)

C₃₂H₂₄Cl₂N₄O₂S (599.5)

C₃₂H₂₂Cl₂N₄OS (581.5)

C₂₄H₂₁Cl₂N₇(478.4)

4

5a

5b

5c

5d

6a

6b

6c

7a

7b

7c

7d

7e

8

9a

9b

10a

10b

10c

10d

11

12

13

14

15

16

17a

17b

18

19a

19b

19c

20a

20b

21

22

23

132.4, 135.5, 135.7, 136.2, 149.7, 158.4 (15 line for 19 sp²car-

bon atoms), 162.2 (CO), 176.4 (CS); its MS, [M^þ], m/z 465

(100%), [M^þþ 2], m/z 467 (45%), [M^þþ 4], m/z 469 (11%).

(CH alkyl), 1696, 1680 (2CO), 1629 (C]N); ¹H NMR

(DMSO-d₆, d, ppm): 1.63e1.67 (m, 2H, CH₂), 2.34 (t, 2H, J ¼

5.86 Hz, CH₂), 2.66 (t, 2H, J ¼ 5.89 Hz, CH₂), 3.00 (s, 2H,

CH₂), 7.03 (d, 2H, J ¼ 8.42 Hz, phenyl), 7.21 (d, 2H, J ¼

8.40 Hz, phenyl), 7.36 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.57 (d,

2H, J ¼ 8.40 Hz, phenyl), 8.18 (s, 1H, CH).

5.1.3. Synthesis of 6-(chlorophenyl)-10-(4-chlorophenyl

methylene)-7,8,9,10-tetrahydrothiazolo

[3⁰,2⁰:1,2]pyrimido[4,5-b]

quinoline-3,5-dione (4)

A mixture of 3 (0.01 mol), chloroacetic acid (0.01 mol) and

anhydrous sodium acetate (0.02 mol) was heated gently with

stirring on a water bath (60 ^ꢃC) for 2 h. The reaction mixture

was allowed to cool to room temperature and poured into

water (100 mL). The deposited precipitate was ﬁltered off

and crystallized from dioxane. The compound was produced

as a yellow powder. IR (KBr, cm^ꢀ1): 3035 (CH aryl), 2917

5.1.4. Synthesis of 2-arylmethylene-6-(4-chlorophenyl)-

10-(4-chlorophenylmethylene)-7,8,9,10-tetrahydro

thiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinoline-3,5-

diones (5aec): general procedure

Method A. A mixture of 3 (0.01 mol), chloroacetic acid

(0.01 mol), the appropriate aromatic aldehyde (0.01 mol)

and (0.02 mol) anhydrous sodium acetate was stirred under

reﬂux in 30 mL of glacial acetic acid and 15 mL of acetic

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

617

anhydride for 15 h. The reaction mixture was cooled and

poured into cold water (100 mL). The deposited precipitate

was ﬁltered off and crystallized from appropriate solvent to

produce 5aec.

Method B. A mixture of 4 (0.01 mol), the appropriate

aromatic aldehyde (0.01 mol) and anhydrous sodium acetate

(0.02 mol) was stirred under reﬂux in 30 mL of glacial acetic

acid and 15 mL of acetic anhydride for 5 h. The reaction

mixture was allowed to cool to room temperature and poured

into cold water (100 mL). The deposited precipitate was

ﬁltered off and crystallized from appropriate solvent to

produce 5a.

(DMSO-d₆, d, ppm): 24.90, 30.72, 30.95 (3C, 3CH₂), 115.6,

119.7, 121.9, 127.1, 127.8, 128.2, 128.8, 129.4, 130.2,

130.5, 133.4, 133.5, 134.8, 136.1, 137.8, 142.0, 143.0,

148.3, 148.7, 152.6, 152.9, 155.6 (22 line for 26 sp²carbon

atoms) and 166.3, 168.6 (2C]O); its MS, [M^þ], m/z 599

(100%), [M^þþ 2], m/z 601 (26%), [M^þþ 4], m/z 603 (6%).

5.1.4.4. Synthesis of 6-(4-chlorophenyl)-10-(4-chlorophenyl-

methylene)-2-(4-N,N-dimethyl-aminophenylmethylene)-7,8,-

9,10-tetrahydrothiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinoline-

3,5-dione (5d). It was obtained from N,N-dimethylaminoben-

zaldehyde as brown crystals and crystallized from dioxane. IR

(KBr, cm^ꢀ1): 3079 (CH aryl), 2924 (CH alkyl), 1700, 1678

1

5.1.4.1. Synthesis of 6-(4-chlorophenyl)-10-(4-chlorophenyl-

methylene)-2-phenylmethylene-7,8,9,10-tetrahydrothiazolo-

[3⁰,2⁰:1,2]pyrimido[4,5-b]quinoline-3,5-dione (5a). It was ob-

tained from benzaldehyde as yellow powder and crystallized

from dioxane. IR (KBr, cm^ꢀ1): 3055 (CH aryl), 2937 (CH al-

kyl), 1687, 1675 (2CO), 1626 (C]N); ¹H NMR (DMSO-d₆, d,

ppm): 1.67 (t, 2H, J ¼ 6.01 Hz, CH₂), 2.35 (t, 2H, J ¼ 5.95 Hz,

CH₂), 2.65e2.82 (m, 2H, CH₂), 6.91e7.10 (m, 3H, phenyl),

7.15e7.26 (m, 2H, phenyl), 7.30e7.41 (m, 2H, phenyl),

7.42e7.72 (m, 4H, phenyl), 8.10 (s, 1H, CH), 8.20 (s, 1H,

CH).

(2CO), 1600 (C]N); H NMR (DMSO-d₆, d, ppm): 1.64 (t,

2H, J ¼ 5.79 Hz, CH₂), 2.33 (t, 2H, J ¼ 5.82 Hz, CH₂),

2.77e2.79 (m, 2H, CH₂), 2.88 (s, 6H, 2CH₃), 7.17 (d, 2H,

J ¼ 8.42 Hz, phenyl), 7.28 (d, 2H, J ¼ 8.40 Hz, phenyl),

7.51e7.72 (m, 4H, phenyl), 7.95 (s, 1H, CH), 8.22 (s, 1H,

CH); its MS, [M^þ], m/z 636 (100%), [M^þþ 2], m/z 638

(56%), [M^þþ 4], m/z 640 (12%).

5.1.5. Synthesis of 3-aryl-11-(4-chlorophenyl)-7-(4-

chlorophenylmethylene)-2,3,7,8,9,10-hexahydroisoxazolo

[5⁰⁰,4⁰⁰:4⁰,5⁰]thiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinolin-12-

ones (6aec): general procedure

5.1.4.2. Synthesis of 6-(4-chlorophenyl)-10-(4-chlorophenyl-

methylene)-2-(4-methoxyphenylmethylene)-7,8,9,10-tetrahy-

drothiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinoline-3,5-dione

(5b). It was obtained from 4-anisaldehyde as yellow powder

and crystallized from dimethylformamide. IR (KBr, cm^ꢀ1):

3051 (CH aryl), 2927 (CH alkyl), 1690, 1676 (2CO), 1632

(C]N); ¹H NMR (DMSO-d₆, d, ppm): 1.65 (t, 2H, J ¼

5.86 Hz, CH₂), 2.34e2.49 (m, 2H, CH₂), 2.81 (t, 2H, J ¼

5.94 Hz, CH₂), 3.81 (s, 3H, OCH₃), 7.10 (d, 2H,

J ¼ 8.42 Hz, phenyl), 7.19 (d, 2H, J ¼ 8.40 Hz, phenyl), 7.47

(d, 2H, J ¼ 8.37 Hz, phenyl), 7.53 (d, 2H, J ¼ 8.39 Hz,

phenyl), 7.58e7.71 (m, 4H, phenyl), 7.86 (s, 1H, CH), 8.07

(s, 1H, CH); ¹³C NMR (DMSO-d₆, d, ppm): 24.9, 30.7, 30.9

(3C, 3CH₂), 55.9 (1C, OCH₃), 114.2, 115.9, 119.7, 127.4,

127.5, 127.8, 127.9, 128.8, 129.4, 130.2, 133.4, 133.5,

133.9, 134.8, 136.1, 142.3, 143.2, 152.6, 152.9, 155.9,

159.9, 163.7 (22 line for 28 sp²carbon atoms) and 166.3,

168.6 (2CO).

A mixture of 5aec (0.01 mol), hydroxylamine hydrochlo-

ride (0.01 mol), and anhydrous sodium acetate (0.01 mol)

was stirred under reﬂux in 30 mL glacial acetic acid for 5 h.

The reaction mixture was allowed to cool to room temperature

and poured into cold water (100 mL). The deposited precipi-

tate was ﬁltered off, dried and crystallized from appropriate

solvent to produce 6aec.

5.1.5.1. Synthesis of 11-(4-chlorophenyl)-7-(4-chlorophenyl-

methylene)-3-phenyl-2,3,7,8,9,10-hexahydroisoxazolo[5⁰⁰,-

4⁰⁰:4⁰,5⁰]thiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinolin-12-

one (6a). It was obtained from 5a as green crystals and crys-

tallized from benzene. IR (KBr, cm^ꢀ1): 3410 (br, NH), 3056

1

(CH aryl), 2928 (CH alkyl), 1692 (CO), 1640 (C]N); H

NMR (DMSO-d₆, d, ppm): 1.65 (t, 2H, J ¼ 5.89 Hz, CH₂)

2.34 (t, 2H, J ¼ 6.00 Hz, CH₂), 2.72e2.83 (m, 2H, CH₂),

6.00 (d, 1H, J ¼ 6.43 Hz, isooxazole proton), 7.02e7.15

(m, 3H, phenyl), 7.18e7.28 (m, 2H, phenyl), 7.30e7.43

(m, 2H, phenyl), 7.52e7.70 (m, 4H, phenyl), 8.24 (s, 1H,

CH), 10.30 (br, NH, D₂O exchangeable); its MS, [M^þ], m/z

608 (100%), [M^þþ 2], m/z 610 (49%), [M^þþ 4], m/z 614

(10%).

5.1.4.3. Synthesis of 6-(4-chlorophenyl)-10-(4-chlorophenyl-

methylene)-2-(2-thienyl-methylene)-7,8,9,10-tetrahydrothiazo-

lo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinoline-3,5-dione (5c). It was

obtained from 2-thiophene carboxaldehyde as brown crystals

and crystallized from dioxane. IR (KBr, cm^ꢀ1): 3023 (CH

5.1.5.2. Synthesis of 11-(4-chlorophenyl)-7-(4-chlorophenyl-

methylene)-3-(4-methoxy-phenyl)-2,3,7,8,9,10-hexahydroisox-

azolo[5⁰⁰,4⁰⁰:4⁰,5⁰]thiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]-quinolin-

12-one (6b). It was obtained from 5b as green powder and

crystallized from ethanol. IR (KBr, cm^ꢀ1): 3400 (br, NH),

3067 (CH aryl), 2923 (CH alkyl), 1685 (CO), 1630 (C]N);

¹H NMR (DMSO-d₆, d, ppm): 1.67 (t, 2H, J ¼ 6.02 Hz,

CH₂), 2.30e2.39 (m, 2H, CH₂), 2.76 (t, 2H, J ¼ 6.00 Hz,

CH₂), 3.87 (s, 3H, OCH₃), 5.89 (d, 1H, J ¼ 6.54 Hz,

1

aryl), 2902 (CH alkyl), 1696, 1672 (2CO), 1651 (C]N); H

NMR (DMSO-d₆, d, ppm): 1.65 (t, 2H, J ¼ 6.00 Hz, CH₂),

2.32 (t, 2H, J ¼ 5.99 Hz, CH₂), 2.72e2.73 (m, 2H, CH₂),

7.21 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.26 (d, 2H, J ¼ 8.40 Hz,

phenyl), 7.31 (t, 1H, J ¼ 4.32 Hz, thiophene), 7.41 (d, 2H,

J ¼ 8.42 Hz, phenyl), 7.55 (d, 2H, J ¼ 8.44 Hz, phenyl), 7.63

(d, 1H, J ¼ 4.92 Hz, thiophene), 7.67 (d, 1H, J ¼ 4.83 Hz,

thiophene), 8.10 (s, 1H, CH), 8.30 (s, 1H, CH); ¹³C NMR

618

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

isooxazole proton), 7.10 (d, 2H, J ¼ 8.43 Hz, phenyl), 7.18 (d,

2H, J ¼ 8.39 Hz, phenyl), 7.45 (d, 2H, J ¼ 8.37 Hz, phenyl),

7.53 (d, 2H, J ¼ 8.39 Hz, phenyl), 7.58e7.76 (m, 4H, phenyl),

8.27 (s, 1H, CH), 10.00 (br s, NH, D₂O exchangeable); its MS,

[M^þ], m/z 638 (100%), [M^þþ 2], m/z 640 (53%), [M^þþ 4],

m/z 642 (15%).

IR (KBr, cm^ꢀ1): 3385 (br, NH), 3061 (CH aryl), 2908 (CH al-

kyl), 1679 (CO), 1635 (C]N); ¹H NMR (DMSO-d₆, d, ppm):

1.36 (t, 3H, J ¼ 7.40 Hz, CH₃), 1.68 (t, 2H, J ¼ 5.69 Hz, CH₂),

2.30 (t, 2H, J ¼ 5.78 Hz, CH₂), 2.70e2.76 (m, 2H, CH₂), 3.48

(q, 2H, J ¼ 7.38 Hz, SCH₂), 7.21 (d, 2H, J ¼ 8.37 Hz, phenyl),

7.34 (d, 2H, J ¼ 8.38 Hz, phenyl), 7.47 (d, 2H, J ¼ 8.41 Hz,

phenyl), 7.57 (d, 2H, J ¼ 8.40 Hz, phenyl), 8.22 (s, 1H, CH),

10.00 (br, NH, D₂O exchangeable); its MS, [M^þ], m/z 493

(100%), [M^þþ 2], m/z 495 (31%), [M^þꢀ CH₃], m/z 478

(36%), [M^þꢀ SC₂H₅], m/z 432 (85%).

5.1.5.3. Synthesis of 11-(4-chlorophenyl)-7-(4-chlorophenyl-

methylene)-3-(2-thienyl)-2,3,7,8,9,10-hexahydroisoxazolo-

[5⁰⁰,4⁰⁰:4⁰,5⁰]thiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinolin-12-

one (6c). It was obtained from 5c as a yellow powder and

crystallized from ethanol. IR (KBr, cm^ꢀ1): 3340 (br, NH),

3049 (CH aryl), 2929 (CH alkyl), 1688 (CO), 1625 (C]N);

¹H NMR (DMSO-d₆, d, ppm): 1.67 (t, 2H, J ¼ 5.97 Hz,

CH₂), 2.33 (t, 2H, J ¼ 5.95 Hz, CH₂), 2.71e2.76 (m, 2H,

CH₂), 6.02 (d, 1H, J ¼ 6.38 Hz, isooxazole proton), 7.21 (d,

2H, J ¼ 8.41 Hz, phenyl), 7.26 (t, 1H, J ¼ 4.40 Hz, thiophene),

7.32 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.46 (d, 2H, J ¼ 8.43 Hz,

phenyl), 7.57 (d, 2H, J ¼ 8.42 Hz, phenyl), 7.67 (d, 1H,

J ¼ 4.86 Hz, thiophene), 7.89 (d, 1H, J ¼ 4.76 Hz, thiophene),

8.30 (s, 1H, CH), 10.80 (br s, NH, D₂O exchangeable); its MS,

[M^þ], m/z 614 (100%), [M^þþ 2], m/z 616 (46%), [M^þþ 4],

m/z 618 (11%).

5.1.6.3. Synthesis of 2-(acetylacetonethio)-5-(4-chlorophenyl)-

9-(4-chlorophenylmethylene)-6,7,8,9-tetrahydropyrimido[4,5-

b]quinolin-4-one (7c). It was obtained from 3 and chloroacety-

lacetone as yellow crystals and crystallized from dioxane. IR

(KBr, cm^ꢀ1): 3410 (br, NH), 3056 (CH aryl), 2918 (CH alkyl),

1720, 1715, 1686, (3CO), 1658 (C]N); ¹H NMR (DMSO-d₆,

d, ppm): 1.74 (t, 2H, J ¼ 5.87 Hz, CH₂), 2.29 (t, 2H,

J ¼ 5.91 Hz, CH₂), 2.69e2.73 (m, 2H, CH₂), 3.11 and 3.16

(2s, 6H, 2COCH₃), 4.04 (s, 1H, CH), 6.94 (d, 2H, J ¼ 8.37 Hz,

phenyl), 7.17 (d, 2H, J ¼ 8.40 Hz, phenyl), 7.27 (d, 2H,

J ¼ 8.39 Hz, phenyl), 7.52 (d, 2H, J ¼ 8.41 Hz, phenyl), 8.32

(s, 1H, CH), 9.80 (br, NH, D₂O exchangeable); ¹³C NMR

(DMSO-d₆, d, ppm): 16.41, 23.06 (2CH₃), 24.21, 27.99, 28.25

(3C, 3CH₂), 40.72 (1C, CH), 115.2, 126.1, 126.9, 129.2, 129.3,

129.5, 129.9, 130.0, 133.7, 135.0, 135.4, 139.1, 139.3, 140.7,

152.3, 154.4 (16 line for 20 sp²carbon atoms), 167.57, 185.68,

192.30 (3CO); its MS, [M^þ], m/z 563 (100%), [M^þþ 2], m/z

565 (46%), [M^þþ 4], m/z 567 (14%), [M^þꢀ COCH₃], m/z

520 (45%), [M^þꢀ CH(COCH₃)₂], m/z 454 (85%), [M^þꢀ

SCH(COCH₃)₂], m/z 422 (19%).

5.1.6. Synthesis of 2-alkylthio-9-(4-chlorophenylmethylene)-

5-(4-chlorophenyl)-6,7,8,9-tetrahydropyrimido[4,5-

b]quinolin-4-ones (7aee): general procedure

To a warmed ethanolic KOH solution (prepared by dis-

solving 0.01 mol of KOH in 50 mL ethanol) was added

each of 3 (0.01 mol), the heating was continued for 30 min

and the mixture was allowed to cool to room temperature,

and the proper halo-compound (0.012 mol) was added. The

mixture was stirred under reﬂux for 5 h, then cooled to

room temperature and poured into cold water (100 mL).

The solid product precipitated was ﬁltered off, washed

with 100 mL water, the product was dried and crystallized

to produce 7aee.

5.1.6.4. Synthesis of 2-(S-acetone)-5-(4-chlorophenyl)-9-(4-

chlorophenylmethylene)-6,7,8,9-tetrahydropyrimido[4,5-b]qui-

nolin-4-one (7d). It was obtained from 3 and chloroacetone as

yellow crystals and crystallized from ethanol. IR (KBr, cm^ꢀ1):

3430 (br, NH), 3059 (CH aryl), 2909 (CH alkyl), 1718, 1683

1

(2CO), 1651 (C]N); H NMR (DMSO-d₆): 1.63e1.66 (m,

5.1.6.1. Synthesis of 5-(4-chlorophenyl)-9-(4-chlorophenyl-

methylene)-2-methylthio-6,7,8,9-tetrahydropyrimido[4,5-b]-

quinolin-4-one (7a). It was obtained from 3 and methyl iodide

(0.012 mol) as pale yellow crystals and crystallized from diox-

ane. IR (KBr, cm^ꢀ1): 3403 (br, NH), 3036 (CH aryl), 2925

2H, CH₂), 1.84 (s, 3H, CH₃), 2.30 (t, 2H, J ¼ 5.98 Hz, CH₂),

2.80 (t, 2H, J ¼ 6.00 Hz, CH₂), 3.98 (s, 2H, CH₂), 7.11 (d,

2H, J ¼ 8.40 Hz, phenyl), 7.20 (d, 2H, J ¼ 8.38 Hz, phenyl),

7.27e7.34 (m, 4H, phenyl), 8.26 (s, 1H, CH), 10.30 (br,

NH, D₂O exchangeable); its MS, [M^þ], m/z 521 (100%),

[M^þþ 2], m/z 523 (52%), [M^þþ 4], m/z 525 (7%),

[M^þꢀ COCH₃], m/z 480 (75%), [M^þꢀ CH₂COCH₃], m/z

466 (66%), [M^þꢀ SCH₂COeCH₃], m/z 434 (28%).

1

(CH alkyl), 1687 (CO), 1652 (C]N); H NMR (DMSO-d₆,

d, ppm): 1.67 (t, 2H, J ¼ 5.96 Hz, CH₂), 2.33 (t, 2H,

J ¼ 6.00 Hz, CH₂), 2.73e2.77 (m, 2H, CH₂), 2.84 (s, 3H,

SCH₃), 7.20 (d, 2H, J ¼ 8.36 Hz, phenyl), 7.33 (d, 2H,

J ¼ 8.45 Hz, phenyl), 7.45 (d, 2H, J ¼ 8.43 Hz, phenyl), 7.59

(d, 2H, J ¼ 8.39 Hz, phenyl), 8.34 (s, 1H, CH), 9.20 (br,

NH, D₂O exchangeable); its MS, [M^þ], m/z 479 (100%),

[M^þþ 2], m/z 481 (56%), [M^þþ 4], m/z 483 (16%)

[M^þꢀ SCH₃], m/z 432 (44%).

5.1.6.5. 2-(S-Phenacyl)-5-(4-chlorophenyl)-9-(4-chlorophenyl-

methylene)-6,7,8,9-tetrahydropyrimido[4,5-b]quinolin-4-one

(7e). It was obtained from 3 and phenacylbromide as pale yel-

low crystals and crystallized from ethanol/dioxane (1:1). IR

(KBr, cm^ꢀ1): 3437 (br, NH), 3078 (CH aryl), 2923 (CH alkyl),

1721, 1689 (2CO), 1647 (C]N); ¹H NMR (DMSO-d₆, d,

ppm): 1.67 (t, 2H, J ¼ 5.96 Hz, CH₂), 2.35 (t, 2H,

J ¼ 5.89 Hz, CH₂), 2.65e2.82 (m, 2H, CH₂), 3.97 (s, 2H,

CH₂), 6.98e7.10 (m, 3H, phenyl), 7.17e7.26 (m, 2H, phenyl),

7.33e7.41 (m, 2H, phenyl), 7.50e7.72 (m, 4H, phenyl), 8.22

5.1.6.2. Synthesis of 5-(4-chlorophenyl)-9-(4-chlorophenyl-

methylene)-2-ethylthio-6,7,8,9-tetrahydropyrimido[4,5-b]qui-

nolin-4-one (7b). It was obtained from 3 and ethyl iodide

(0.012 mol) as orange crystals and crystallized from dioxane.

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

619

(s, 1H, CH), 10.30 (br, NH, D₂O exchangeable); its MS, [M^þ],

m/z 583 (100%), [M^þþ 2], m/z 585 (53%), [M^þþ 4], m/z 587

(10%), [M^þꢀ C₆H₅], m/z 506 (100%), [M^þꢀ COC₆H₅], m/z

478 (15%).

[M^þ], m/z 503 (100%), [M^þþ 2], m/z 505 (55%), [M^þþ 4],

m/z 507 (13%), [M^þꢀ CH₃], m/z 488 (53%).

5.1.8.2. Synthesis of 6-(4-chlorophenyl)-10-(4-chlorophenyl-

methylene)-3-phenyl-7,8,9,10-tetrahydrothiazolo[3⁰,2⁰:1,2]-

pyrimido[4,5-b]quinolin-5-one (9b). It was obtained from 7e

as yellow crystals and crystallized from dimethylformamide.

IR (KBr, cm^ꢀ1): 3054 (CH aryl), 2918 (CH alkyl), 1695

5.1.7. Synthesis of 5-(4-chlorophenyl)-9-

(4-chlorophenylmethylene)-2-hydrazino-6,7,8,9-

tetrahydropyrimido[4,5-b]quinolin-4-one (8)

1

(CO), 1647 (C]N); H NMR (DMSO-d₆, d, ppm): 1.66e

A suspension of 7a,b (0.01 mol) in hydrazine hydrate

(99e100%) (25 mL) was stirred under gentle reﬂux. The

insoluble solid went into solution within 10 min with copious

evolution of methylmercaptan to form a clear solution. After

30 min when the solid product started separating out, heating

was continued for 8 h and the reaction mixture was allowed to

cool to room temperature. The solid which separated was

ﬁltered, washed with ethanol and dried, and obtained as yellow

powder crystals and crystallized from dimethylformamide. IR

(KBr, cm^ꢀ1): 3198 (br s, NH), 3036 (CH aryl), 2934 (CH al-

kyl), 1676 (CO), 1676 (C]N); ¹H NMR (DMSO-d₆, d, ppm):

1.64e1.69 (m, 2H, CH₂), 2.31 (t, 2H, J ¼ 5.98 Hz, CH₂), 2.77

(t, 2H, J ¼ 6.01 Hz, CH₂), 7.09 (d, 2H, J ¼ 8.34 Hz, phenyl),

7.40 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.43e7.47 (m, 4H, phenyl),

8.02 (s, 1H, CH), 8.75 (br s, NH₂, D₂O exchangeable), 9.50,

11.00 (2 br s, 2NH, D₂O exchangeable); ¹³C NMR (DMSO-

d₆, d, ppm): 22.27, 22.73, 27.19 (3C, 3CH₂), 109.1, 127.7,

127.8, 128.3, 129.3, 129.4, 131.1, 131.6, 133.4, 136.1,

136.3, 137.9, 141.5, 148.2 150.0, 155.1 (16 lines for 20 sp²

carbon atoms), 160.2 (CO); its MS, [M^þ], m/z 463 (100%),

[M^þþ 2], m/z 465 (56%), [M^þþ 4], m/z 467 (14%).

1.70 (m, 2H, CH₂), 2.28 (t, 2H, J ¼ 5.93 Hz, CH₂), 2.75 (t,

2H, J ¼ 5.99 Hz, CH₂), 6.97e7.05 (m, 4H, phenyl), 7.22 (d,

2H, J ¼ 8.39 Hz, phenyl), 7.46e7.80 [m, 6H (5H, phenyl þ s,

1H, thiazole)], 8.08 (d, 2H, J ¼ 8.43 Hz, phenyl), 8.28 (s, 1H,

CH); its MS, [M^þ], m/z 565 (100%), [M^þþ 2], m/z 567

(57%), [M^þþ 4], m/z 569 (12%), [M^þꢀ C₆H₅], m/z 488

(41%).

5.1.9. Synthesis of 3-alkyl-2-alkylthio-5-(4-chlorophenyl)-9-

(4-chlorophenylmethylene)-6,7,8,9-tetrahydropyrimido[4,5-

b]quinolin-4-ones (10aed): general procedure

To a warmed ethanolic sodium ethoxide solution (prepared

by dissolving 0.01 mol of sodium metal in 30 mL absolute eth-

anol) was added each of 7a or 7b (0.01 mol), the heating was

continued for 30 min, and the mixture was allowed to cool to

room temperature and the proper alkyliodide (0.012 mol) was

added. The mixture was stirred under reﬂux for 3 h, cooled to

room temperature, and poured into cold water (100 mL). The

solid so-precipitated was ﬁltered off, washed with water and

dried, to produce 10aec in high yields.

5.1.8. Synthesis of 6-(4-chlorophenyl)-10-(4-

chlorophenylmethylene)-3-(methyl or phenyl)-7,8,9-

hexahydrothiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinolin-5-

ones (9a,b): general procedure

A solution of 7d,e (0.01 mol) in glacial acetic acid (40 mL)

and catalytic amount of sulfuric acid (1 mL) was stirred under

reﬂux for 8 h. The reaction mixture was allowed to cool,

poured into cold water (100 mL), neutralized by ammonia

solution, and the solid precipitate was ﬁltered off, washed

with water, dried and crystallized to produce 9a,b.

5.1.9.1. Synthesis of 5-(4-chlorophenyl)-2-methylthio-3-methyl-

9-(4-chlorophenylmethylene)-6,7,8,9-tetrahydropyrimido[4,5-

b]quinolin-4-one (10a). It was obtained from 7a and methyl

iodode as red crystals and crystallized from benzene. IR

(KBr, cm^ꢀ1): 3056 (CH aryl), 2940 (CH alkyl), 1696 (CO),

1618 (C]N); ¹H NMR (CDCl₃, d, ppm): 1.64e1.68 (m,

2H, CH₂), 2.24 (t, 2H, J ¼ 5.90 Hz, CH₂), 2.65 (s, 3H,

SCH₃), 2.76 (t, 2H, J ¼ 5.93 Hz, CH₂), 4.20 (s, 3H, Ne

CH₃), 6.97 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.08 (d, 2H,

J ¼ 8.40 Hz, phenyl), 7.21 (d, 2H, J ¼ 8.43 Hz, phenyl), 7.51

(d, 2H, J ¼ 8.38 Hz, phenyl), 8.24 (s, 1H, CH); its MS,

[M^þ], m/z 493 (100%), [M^þþ 2], m/z 495 (61%),

[M^þꢀ CH₃], m/z 478 (12%), [M^þꢀ SCH₃], m/z 446 (5%).

5.1.8.1. Synthesis of 6-(4-chlorophenyl)-10-(4-chlorophenyl-

methylene)-3-methyl-7,8,9,10-tetrahydrothiazolo[3⁰,2⁰:1,2]-

pyrimido[4,5-b]quinolin-5-one (9a). The compound was ob-

tained from 7d as yellow crystals and crystallized from dime-

thylformamide. IR (KBr, cm^ꢀ1): 3066 (CH aryl), 2908 (CH

alkyl), 1696 (CO), 1650 (C]N); ¹H NMR (DMSO-d₆, d,

ppm): 1.65e1.69 (m, 2H, CH₂), 2.13 (s, 3H, CH₃), 2.30 (t,

5.1.9.2. Synthesis of 5-(4-chlorophenyl)-3-ethyl-2-methylthio-

9-(4-chlorophenylmethylene)-6,7,8,9-tetrahydropyrimido[4,5-

b]quinolin-4-one (10b). It was obtained from 7a and ethyl

iodode as yellow powder and crystallized from dioxane. IR

(KBr, cm^ꢀ1): 3075 (CH aryl), 2920 (CH alkyl), 1705 (CO),

1625 (C]N); ¹H NMR (CDCl₃, d, ppm): 1.30 (t, 3H, J

¼ 7.38 Hz, CH₃), 1.66e1.69 (m, 2H, CH₂), 2.27 (t, 2H, J ¼

5.86 Hz, CH₂), 2.68 (s, 3H, SCH₃), 2.78 (t, 2H, J ¼ 5.90 Hz,

CH₂), 4.38 (q, 2H, J ¼ 7.49 Hz, NeCH₂), 6.98 (d, 2H,

J ¼ 8.35 Hz, phenyl), 7.10 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.25

(d, 2H, J ¼ 8.39 Hz, phenyl), 7.55 (d, 2H, J ¼ 8.38 Hz,

2H,

J ¼ 5.85 Hz,

CH₂),

2.76

(t,

2H,

J ¼

5.81 Hz, CH₂), 7.13 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.39 (d,

2H, J ¼ 8.36 Hz, phenyl), 7.42e7.57 (m, 4H, phenyl), 7.83

(s, 1H, thiazole), 8.30 (s, 1H, CH); ¹³C NMR (DMSO-d₆, d,

ppm): 21.56 (CH₃), 25.71, 29.83, 30.09 (3C, 3CH₂), 106.7,

118.9, 127.4, 126.5, 128.3, 128.7, 129.2, 131.4, 133.6,

134.2, 135.4, 136.7, 142.5, 145.5, 148.4, 152.1, 152.8, 154.2

(18 line for 22 sp²carbon atoms), 166.7 (CO); its MS,

620

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

phenyl), 8.18 (s, 1H, CH); its MS, [M^þ], m/z 507 (63%),

[M^þꢀ C₂H₅], m/z 478 (18%), [M^þꢀ SC₂H₅], m/z 446 (10%).

(100 mL). The deposited precipitate was ﬁltered off, dried,

and obtained as pale yellow powder and crystallized from eth-

anol/dioxane (1:1). IR (KBr, cm^ꢀ1): 3400 (br s, NH’s), 3023

1

5.1.9.3. Synthesis of 5-(4-chlorophenyl)-2-ethylthio-3-methyl-

9-(4-chlorophenylmethylene)-6,7,8,9-tetrahydropyrimido[4,5-

b]quinolin-4-one (10c). It was obtained from 7b and methyl

iodode as red crystals and crystallized from benzene. IR

(KBr, cm^ꢀ1): 3065 (CH aryl), 2930 (CH alkyl), 1698 (CO),

1620 (C]N); ¹H NMR (CDCl₃, d, ppm): 1.21 (t, 3H,

J ¼ 7.36 Hz, CH₃), 1.68e1.71 (m, 2H, CH₂), 2.29 (t, 2H,

J ¼ 5.91 Hz, CH₂), 2.73 (q, 2H, J ¼ 7.35 Hz, SCH₂), 2.79 (t,

2H, J ¼ 5.87 Hz, CH₂), 4.32e4.40 (s, 3H, NeCH₃), 6.97 (d,

2H, J ¼ 8.34 Hz, phenyl), 7.11 (d, 2H, J ¼ 8.38 Hz, phenyl),

7.28 (d, 2H, J ¼ 8.38 Hz, phenyl), 7.58 (d, 2H, J ¼ 8.39 Hz,

phenyl), 8.27 (s, 1H, CH); its MS, [M^þ], m/z 507 (88%),

[M^þþ 2], m/z 509 (51%), [M^þþ 4], m/z 511 (10%).

(CH aryl), 2929 (CH alkyl), 1687 (CO), 1653 (C]N); H

NMR (DMSO-d₆, d, ppm): 1.68 (t, 2H, J ¼ 5.98 Hz, CH₂),

2.27 (t, 2H, J ¼ 5.92 Hz, CH₂), 2.41 (s, 3H, CH₃), 2.44 (s,

3H, CH₃), 2.70e2.75 (m, 2H, CH₂), 6.95e6.99 (m, 4H,

phenyl), 7.10 (d, 2H, J ¼ 8.39 Hz, phenyl), 7.34 (d, 2H,

J ¼ 8.40 Hz, phenyl), 8.25 (s, 1H, CH), 9.34, 10.10 (2 br s,

2NH, D₂O exchangeable); its MS, [M^þ], m/z 559 (100%),

[M^þþ 2], m/z 561 (54%), [M^þþ 4], m/z 563 (21%).

5.1.12. Synthesis of 2-(4,6-dimethyl-2-thioxo-1,2-

dihydropyrimidin-5-ylthio)-9-(4-chloro-phenylmethylene)-

5-(4-chlorophenyl)-6,7,8,9-tetrahydropyrimido[4,5-b]

quinolin-4-one (13)

A mixture of 7c (0.01 mol) and thiourea (0.01 mol) was

stirred under reﬂux in dioxane (30 mL) in the presence of

catalytic amount of piperidine for 15 h. The reaction mixture

was allowed to cool to room temperature, poured into water

(100 mL), the deposited precipitate was ﬁltered off, washed

with ethanol (30 mL), dried and crystallized from dimethylfor-

mamide as yellow powder. IR (KBr, cm^ꢀ1): 3420 (br s, NH’s),

3043 (CH aryl), 2928 (CH alkyl), 1686, (CO), 1630 (C]N);

¹H NMR (DMSO-d₆, d, ppm): 1.67 (t, 2H, J ¼ 5.69 Hz,

CH₂), 2.27 (t, 2H, J ¼ 5.74 Hz, CH₂), 2.45 (s, 3H, CH₃),

2.52 (s, 3H, CH₃), 2.72e2.77 (m, 2H, CH₂), 7.05 (d, 2H,

J ¼ 8.34 Hz, phenyl), 7.15 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.34

(d, 2H, J ¼ 8.43 Hz, phenyl), 7.54 (d, 2H, J ¼ 8.43 Hz,

phenyl), 8.30 (s, 1H, CH), 9.20, 10.50 (2 br s, 2NH, D₂O

exchangeable); its MS, [M^þ], m/z 603 (53%), [M^þþ 2], m/z

605 (34%), [M^þþ 4], m/z 607 (18%).

5.1.9.4. Synthesis of 5-(4-chlorophenyl)-3-ethyl-2-ethylthio-9-

(4-chlorophenylmethylene)-6,7,8,9-tetrahydropyrimido[4,5-

b]quinolin-4-one (10d). It was obtained from 7b and ethyl

iodode as yellow powder and crystallized from dioxane. IR

(KBr, cm^ꢀ1): 3029 (CH aryl), 2921 (CH alkyl), 1700 (CO),

1

1650 (C]N); H NMR (CDCl₃, d, ppm): 1.15e1.20 (t, 3H,

CH₃), 1.29 (t, 3H, J ¼ 7.43 Hz, CH₃), 1.64e1.70 (m, 2H,

CH₂), 2.30 (t, 2H, J ¼ 5.89 Hz, CH₂), 2.70 (q, 2H,

J ¼ 7.56 Hz, SCH₂), 2.76e2.81 (t, 2H, CH₂), 4.26e4.30 (q,

2H, NeCH₂), 7.05 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.15 (d,

2H, J ¼ 8.38 Hz, phenyl), 7.30 (d, 2H, J ¼ 8.36 Hz, phenyl),

7.65 (d, 2H, J ¼ 8.41 Hz, phenyl), 8.22 (s, 1H, CH); its MS,

[M^þ], m/z 521 (100%), [M^þþ 2], m/z 523 (60%), [M^þþ 4],

m/z 525 (19%).

5.1.10. Synthesis of 5-(4-chlorophenyl)-2-methylsulfone-9-

(4-chlorophenylmethylene)-6,7,8,9-tetrahydropyrimido[4,5-

b]quinolin-4-one (11)

5.1.13. Synthesis of 2-acetyl-6-(4-chlorophenyl)-10-(4-

chlorophenylmethylene)-3-methyl-7,8,9,10-

tetrahydrothiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]

quinolin-5-one (14)

A mixture of 7a (0.01 mol) and excess amount of hydrogen

peroxide (5 mL) in acetic acid (30 mL) was heated gently with

stirring for 10 h. The reaction mixture was allowed to cool to

0 ^ꢃC. The deposited precipitate was ﬁltered off and crystal-

lized from dioxane. IR (KBr, cm^ꢀ1): 3385 (br, NH), 3031

(CH aryl), 2909 (CH alkyl), 1635 (C]N), 1165, 1345

A solution of 7c (0.01 mol) in a mixture of acetic anhy-

dride/pyridine (20:10) was stirred under reﬂux for 4 h. The

reaction mixture was allowed to cool to room temperature,

poured into cold water (100 mL), the deposited precipitate

was ﬁltered off, dried and crystallized from benzene. The com-

pound was obtained as yellow crystals. IR (KBr, cm^ꢀ1): 3066

(CH aryl), 2927 (CH alkyl), 1720, 1687, (2CO), 1630 (C]N);

¹H NMR (DMSO-d₆, d, ppm): 1.68 (t, 2H, J ¼ 6.01 Hz, CH₂),

2.28 (t, 2H, J ¼ 6.03 Hz, CH₂), 2.45 (s, 3H, CH₃), 2.70e2.75

(m, 2H, CH₂), 2.83 (s, 3H, CH₃), 6.97e7.12 (m, 4H, phenyl),

7.24 (d, 2H, J ¼ 8.41 Hz, phenyl), 7.54 (d, 2H, J ¼ 8.43 Hz,

phenyl), 8.25 (s, 1H, CH); ¹³C NMR (DMSO-d₆, d, ppm):

12.70, 24.81 (2C, 2CH₃), 25.72, 29.91, 30.13 (3C, 3CH₂),

104.2, 119.8, 127.8, 127.9, 128.8, 128.9, 129.4, 130.3,

133.5, 133.6, 134.9, 136.1, 143.1, 145.2, 148.6, 152.7,

152.9, 155.1 (18 line for 22 sp²carbon atoms), 165.3, 193.1

(2CO); its MS, [M^þ], m/z 545 (100%), [M^þþ 2], m/z 547

(56%), [M^þþ 4], m/z 549 (11%), [M^þꢀ CH₃], m/z 530

(15%), [M^þꢀ COCH₃], m/z 502 (34%).

1

(SO₂); H NMR (DMSO-d₆, d, ppm): 1.69 (t, 2H, J ¼ 5.86

Hz, CH₂), 2.29 (t, 2H, J ¼ 5.69 Hz, CH₂), 2.71e2.76 (m,

2H, CH₂), 3.30 (s, 3H, SO₂CH₃), 7.18 (d, 2H, J ¼ 8.40 Hz,

phenyl), 7.35 (d, 2H, J ¼ 8.39 Hz, phenyl), 7.55 (d, 2H,

J ¼ 8.38 Hz, phenyl), 7.68 (d, 2H, J ¼ 8.37 Hz, phenyl), 8.30

(s, 1H, CH), 9.80 (br, NH, D₂O exchangeable); its MS,

[M^þ], m/z 511 (100%), [M^þþ 2], m/z 513 (63%), [M^þþ 4],

m/z 515 (16%).

5.1.11. Synthesis of 2-[3,5-dimethyl-1,2-dihydropyrazol-4-

ylthio]-9-(4-chlorophenyl)-5-(4-chlorophenylmethylene)-

6,7,8,9-tetrahydropyrimido[4,5-b]quinolin-4-one (12)

A mixture of 7c (0.01 mol) and hydrazine hydrate (99e

100%) in dioxane (20 mL) and ethanol (10 mL) was stirred

under reﬂux for 12 h. The reaction mixture was allowed to

cool to room temperature and poured into cold water

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

621

5.1.14. Synthesis of 2-(acetoxime)-6-(4-chlorophenyl)-

10-(4-chlorophenylmethylene)-3-methyl-7,8,9,10-

from dioxane. IR (KBr, cm^ꢀ1): 3054 (CH aryl), 2917 (CH alkyl),

1703, 1685, (2CO), 1615 (C]N); ¹H NMR (DMSO-d₆, d, ppm):

1.65e1.71 (m, 2H, CH₂), 2.18 (s, 3H, CH₃), 2.27e2.33 (m, 2H,

CH₂), 2.68e2.73 (m, 2H, CH₂), 5.28, 5.50 (2d, 2H, CH]CH,

J ¼ 11.5 Hz), 7.01 (d, 2H, J ¼ 8.40 Hz, phenyl), 7.10e7.26 (m,

4H, phenyl), 7.27e7.34 (m, 5H, phenyl), 7.58 (d, 2H,

J ¼ 8.42 Hz, phenyl), 8.34 (s, 1H, CH); its MS, [M^þ], m/z 633

(100%), [M^þþ 2], m/z 635 (52%), [M^þþ 4], m/z 637 (26%).

tetrahydrothiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]

quinolin-5-one (15)

A mixture of 14 (0.01 mol), hydroxylamine hydrochloride

(0.01 mol) in dioxane (30 mL) and catalytic amount of piper-

idine was added. The reaction mixture was stirred under reﬂux

for 15 h, allowed to cool to room temperature, and poured into

water (100 mL). The deposited precipitate was ﬁltered off,

dried and crystallized from ethanol/dioxane (1:1). IR (KBr,

cm^ꢀ1): 3540 (br s, OH), 3045 (CH aryl), 2923 (CH alkyl),

1690 (CO), 1650 (C]N); ¹H NMR (DMSO-d₆, d, ppm):

1.70 (t, 2H, J ¼ 5.97 Hz, CH₂), 2.30 (t, 2H, J ¼ 5.95 Hz,

CH₂), 2.41 (s, 3H, CH₃), 2.73e2.78 (m, 2H, CH₂), 3.02 (s,

3H, CH₃), 3.47 (br, OH overlapped with H₂O of DMSO,

D₂O exchangeable), 7.04 (d, 2H, J ¼ 8.36 Hz, phenyl), 7.20

(d, 2H, J ¼ 8.40 Hz, phenyl), 7.35 (d, 2H, J ¼ 8.40 Hz, phe-

nyl), 7.54 (d, 2H, J ¼ 8.37 Hz, phenyl), 8.25 (s, 1H, CH); its

MS, [M^þ], m/z 560 (47%), [M^þþ 2], m/z 562 (26%),

[M^þþ 4], m/z 564 (6%), [M^þꢀ OH], m/z 543 (100%).

5.1.16.2. Synthesis of 6-(4-chlorophenyl)-2-E-(4-methoxy-cin-

namoyl)-10-(4-chlorophenylmethylene)-3-methyl-7,8,9,10-tet-

rahydrothiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinolin-5-one

(17b). It was obtained from 4-anisaldehyde as brown powder

and crystallized from dioxane. IR (KBr, cm^ꢀ1): 3045 (CH

1

aryl), 2931 (CH alkyl), 1700, 1685 (2CO), 1620 (C]N); H

NMR (DMSO-d₆, d, ppm): 1.66e1.72 (m, 2H, CH₂), 2.21

(s, 3H, CH₃), 2.26e2.31 (m, 2H, CH₂), 2.71e2.78 (m, 2H,

CH₂), 3.86 (s, 3H, OCH₃), 5.30, 5.53 (2d, 2H, CH]CH,

J ¼ 10.9 Hz), 6.98 (d, 2H, J ¼ 8.38 Hz, phenyl), 7.13e7.29

(m, 4H, phenyl), 7.23e7.44 (m, 4H, phenyl), 7.59 (d, 2H,

J ¼ 8.37 Hz, phenyl), 8.50 (s, 1H, CH); its MS, [M^þ], m/z

663 (78%), [M^þþ 2], m/z 665 (29%).

5.1.15. Synthesis of 2-(acetothiosemicarbazone)-6-

(4-chlorophenyl)-10-(4-chlorophenylmethylene)-3-

methyl-7,8,9,10-tetrahydrothiazolo[3⁰,2⁰:1,2]pyrimido

[4,5-b]quinolin-5-one (16)

5.1.17. Synthesis of 4-chloro-5-(4-chlorophenyl)-9-

(4-chlorophenylmethylene)-6,7,8,9-tetrahydropyrimido

[4,5-b]quinoline (18)

A mixture of 14 (0.01 mol), thiosemicarbazide (0.01 mol)

in dioxane (30 mL) and catalytic amount of piperidine was

added. The reaction mixture was stirred under reﬂux for

12 h, allowed to cool to room temperature, and poured into

water (100 mL). The deposited precipitate was ﬁltered off,

dried and crystallized from dioxane. The compound was

obtained as green powder. IR (KBr, cm^ꢀ1): 3400 (br s, NH),

3071 (CH aryl), 2918 (CH alkyl), 1689, (CO), 1625 (C]N);

¹H NMR (DMSO-d₆, d, ppm): 1.66 (t, 2H, J ¼ 6.00 Hz,

CH₂), 2.31 (t, 2H, J ¼ 6.02 Hz, CH₂), 2.40 (s, 3H, CH₃),

2.74e2.79 (m, 2H, CH₂), 2.96 (s, 3H, CH₃), 7.05 (d, 2H,

J ¼ 8.36 Hz, phenyl), 7.22 (d, 2H, J ¼ 8.38 Hz, phenyl), 7.2

(d, 2H, J ¼ 8.39 Hz, phenyl), 7.55 (d, 2H, J ¼ 8.36 Hz, phe-

nyl), 8.22 (s, 1H, CH), 8.67 (br, NH₂, D₂O exchangeable),

9.80 (br, NH, D₂O exchangeable); its MS, [M^þ], m/z 618

(47%), [M^þþ 2], m/z 620 (21%), [M^þþ 4], m/z 622 (5%),

[M^þꢀ CH₃], m/z 603 (53%).

A solution of 7a (0.01 mol) in dry dioxane (30 mL) was

treated with 7 mL of phosphorus oxychloride and the mixture

was stirred under reﬂux for 3 h. The reaction mixture was

allowed to cool to room temperature, poured into cold water

(100 mL) whereby a solid was separated, ﬁltered off and crystal-

lized from dioxane (yellow powder) in 56% yield, m.p.

313e315 ^ꢃC (melted); IR (KBr, cm^ꢀ1): 3049 (CH aryl), 2919

1

(CH alkyl), 1600 (C]N), 1160 (CeCl); H NMR (CDCl₃, d,

ppm): 1.67 (t, 2H, J ¼ 6.00 Hz, CH₂), 2.30 (t, 2H, J ¼ 5.96 Hz,

CH₂), 2.65e2.72 (m, 2H, CH₂), 2.84 (s, 3H, SCH₃), 6.96 (d,

2H, J ¼ 8.39 Hz, phenyl), 7.20 (d, 2H, J ¼ 8.38 Hz, phenyl),

7.37 (d, 2H, J ¼ 8.40 Hz, phenyl), 7.47 (d, 2H, J ¼ 8.41 Hz, phe-

nyl), 8.25 (s, 1H, CH); its MS, [M^þ], m/z 497 (100%), [M^þþ 2],

m/z 499 (59%), [M^þþ 4], m/z 501 (17%).

5.1.18. Synthesis of 4-arylamino-5-(4-chlorophenyl)-

9-(chlorophenylmethylene)-2-methylthio-6,7,8,9-

5.1.16. Synthesis of 2-E-cinnamoyl-6-(4-chlorophenyl)-10-

(4-chlorophenylmethylene)-3-methyl-7,8,9,10-

tetrahydropyrimido[4,5-b]quinoline

(19aec or 20a,b): general procedure

tetrarhydrothiazolo[3⁰,2⁰:1,2]pyrimido[4,5-b]

In a warm solution of 18 (0.01 mol) in glacial acetic acid

(40 mL) was added the freshly distilled aryl-amine or 2nd amine

(0.01 mol). The reaction mixture was stirred under reﬂux for 3 h,

then allowed to cool to 0 ^ꢃC for 4 h, and the solid obtained was

ﬁltered, washed with water (100 mL) dried and recrystallized

from appropriate solvent to produce 19aec and 20a,b.

quinolin-5-one (17a,b): general procedure

A mixture of 14 (0.01 mol), aldehyde (0.01 mol) and a cata-

lytic amount of piperidine was heated at 170e180 ^ꢃC in test

tube for 3 h. The product was solidiﬁed by cooling and addition

of methanol (50 mL). The precipitate formed was collected by

ﬁltration and crystallized from an appropriate solvent.

5.1.18.1. Synthesis of 5-(4-chlorophenyl)-9-(chlorophenyl-

methylene)-4-phenylamino-2-methylthio-6,7,8,9-tetrahydropyr-

imido[4,5-b]quinoline (19a). It was obtained from aniline as

pale yellow crystals and crystallized from dioxane. IR (KBr,

cm^ꢀ1): 3410 (br s, NH), 3032 (CH aryl), 2919 (CH alkyl),

5.1.16.1. Synthesis of 2-E-cinnamoyl-6-(4-chlorophenyl)-10-

(4-chlorophenylmethylene)-3-methyl-7,8,9,10-tetrahydrothia

zolo[3⁰,2⁰:1,2]pyrimido[4,5-b]quinolin-5-one (17a). It was ob-

tained from benzaldehyde as brown powder and crystallized

622

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

1609 (C]N); ¹H NMR (DMSO-d₆, d, ppm): 1.66 (t, 2H,

J ¼ 5.97 Hz, CH₂), 2.31 (t, 2H, J ¼ 5.89 Hz, CH₂),

2.67e2.72 (m, 2H, CH₂), 2.86 (s, 3H, SCH₃), 6.93e7.12 (m,

3H, phenyl), 7.22 (d, 2H, J ¼ 8.43 Hz, phenyl), 7.36e7.42

(m, 2H, phenyl), 7.45e7.53 (m, 4H, phenyl), 7.65 (d, 2H,

J ¼ 8.41 Hz, phenyl), 8.35 (s, 1H, CH), 9.20 (br, NH, D₂O ex-

changeable); its MS, [M^þ], m/z 554 (73%), [M^þþ 2], m/z 556

(29%), [M^þþ 4], m/z 558 (7%).

as yellow crystals and crystallized from dimethylformamide.

IR (KBr, cm^ꢀ1): 3415 (br s, NH), 3067 (CH aryl), 2951 (CH

alkyl), 1630 (C]N); H NMR (DMSO-d₆, d, ppm): 1.71 (t,

1

2H, J ¼ 6.02 Hz, CH₂), 2.28 (t, 2H, J ¼ 6.00 Hz, CH₂),

2.67e2.72 (m, 2H, CH₂), 2.77 (s, 3H, SCH₃), 3.26e3.31 (m,

2H, CH₂), 3.39e3.43 (m, 2H, CH₂), 3.82e3.90 (m, 4H,

2CH₂), 7.04 (d, 2H, J ¼ 8.38 Hz, phenyl), 7.26 (d, 2H, J ¼

8.37 Hz, phenyl), 7.39 (d, 2H, J ¼ 8.40 Hz, phenyl), 7.57 (d,

2H, J ¼ 8.41 Hz, phenyl), 8.33 (s, 1H, CH); its MS, [M^þ],

m/z 544 (65%), [M^þþ 2], m/z 546 (31%), [M^þþ 4], m/z

548 (8%).

5.1.18.2. Synthesis of 4-(chlorophenylamino)-5-(4-chloro-

phenyl)-9-(chlorophenylmethylene)-2-methylthio-6,7,8,9-tet-

rahydropyrimido[4,5-b]quinoline (19b). It was obtained from

4-chloroaniline as yellow crystals and crystallized from dime-

thylformamide. IR (KBr, cm^ꢀ1): 3395 (br s, NH), 3049 (CH

5.1.19. Synthesis of 4-(o-carboxyphenyl)amino-5-

(4-chlorophenyl)-9-(chlorophenylmethylene)-2-methylthio-

6,7,8,9-tetrahydropyrimido[4,5-b]quinoline (21)

1

aryl), 2926 (CH alkyl), 1617 (C]N); H NMR (DMSO-d₆,

d, ppm): 1.67 (t, 2H, J ¼ 5.86 Hz, CH₂), 2.30 (t, 2H,

J ¼ 5.88 Hz, CH₂), 2.68e2.74 (m, 2H, CH₂), 2.88 (s, 3H,

SCH₃), 6.98e7.18 (m, 4H, phenyl), 7.23 (d, 2H, J ¼ 8.44

Hz, phenyl), 7.34e7.43 (m, 4H, phenyl), 7.65 (d, 2H,

J ¼ 8.42 Hz, phenyl), 8.32 (s, 1H, CH), 9.50 (br, NH, D₂O

exchangeable); its MS, [M^þ], m/z 588 (100%), [M^þþ 2], m/z

590 (57%), [M^þþ 4], m/z 592 (10%).

It was obtained from anthranilic acid as yellow powder and

crystallized from dioxane. IR (KBr, cm^ꢀ1): 3510 (br, OH),

3368 (br, NH), 3041 (CH aryl), 2927 (CH alkyl), 1716

1

(CO), 1605 (C]N); H NMR (DMSO-d₆, d, ppm): 1.68 (t,

2H, J ¼ 5.86 Hz, CH₂), 2.27 (t, 2H, J ¼ 5.90 Hz, CH₂),

2.69e2.76 (m, 2H, CH₂), 2.85 (s, 3H, SCH₃), 6.92e7.15 (m,

4H, phenyl), 7.22 (d, 2H, J ¼ 8.34 Hz, phenyl), 7.34e7.40

(m, 2H, phenyl), 7.49 (d, 1H, J ¼ 9.63 Hz, phenyl), 7.57 (d,

2H, J ¼ 8.32 Hz, phenyl), 7.70 (d, 1H, J ¼ 9.46 Hz, phenyl),

8.25 (s, 1H, CH), 9.10 (br, NH, D₂O exchangeable), 12.30

(br s, OH, D₂O exchangeable); its MS, [M^þ], m/z 598

(100%), [M^þþ 2], m/z 600 (67%), [M^þþ 4], m/z 602 (17%).

5.1.18.3. Synthesis of 5-(4-chlorophenyl)-9-(chlorophenyl-

methylene)-4-(4-methoxyphenyl-amino)-2-methylthio-6,7,8,9-

tetrahydropyrimido[4,5-b]quinoline (19c). It was obtained

from 4-anisidene as yellow crystals and crystallized from

dimethylformamide in 76% yield, m.p. 270e272 ^ꢃC (melted);

IR (KBr, cm^ꢀ1): 3420 (br s, NH), 3054 (CH aryl), 2926 (CH

5.1.20. Synthesis of 14-(4-chlorophenyl)-10-

1

alkyl), 1610 (C]N); H NMR (DMSO-d₆, d, ppm): 1.65 (t,

(4-chlorophenylmethylene)-7-methylthio-10,11,12,13-

tetrahydro-5H-quino[2⁰,3⁰:4,5]pyrimido[6,1-b]

quinazolin-5-one (22)

2H, J ¼ 5.96 Hz, CH₂), 2.30 (t, 2H, J ¼ 5.99 Hz, CH₂),

2.67e2.72 (m, 2H, CH₂), 2.83 (s, 3H, SCH₃), 3.89 (s, 3H,

OCH₃), 6.92e7.16 (m, 4H, phenyl), 7.25 (d, 2H, J ¼

8.38 Hz, phenyl), 7.33 (d, 2H, J ¼ 8.39 Hz, phenyl), 7.45 (d,

2H, J ¼ 8.37 Hz, phenyl), 7.65 (d, 2H, J ¼ 8.40 Hz, phenyl),

8.40 (s, 1H, CH), 10.00 (br, NH, D₂O exchangeable); its

MS, [M^þ], m/z 584 (100%), [M^þþ 2], m/z 586 (59%),

[M^þþ 4], m/z 588 (22%).

A solution of 21 (0.01 mol) in glacial acetic acid (40 mL)

and catalytic amount of sulfuric acid (1 mL) was stirred under

reﬂux for 8 h. The reaction mixture was allowed to cool,

poured into cold water (100 mL), neutralized by ammonia

solution, the solid so-precipitated was ﬁltered off, washed

with water, dried and crystallized from dimethylformamide

as yellow crystals. IR (KBr, cm^ꢀ1): 3023 (CH aryl), 2917

1

5.1.18.4. Synthesis of 5-(4-chlorophenyl)-9-(chlorophenyl

methylene)-4-piprazino-2-methylthio-6,7,8,9-tetrahydropyri-

mido[4,5-b]quinoline (20a). It was obtained from piperazine

as pale yellow crystals and crystallized from dioxane. IR

(KBr, cm^ꢀ1): 3365 (br, NH), 3061 (CH aryl), 2931 (CH alkyl),

1600 (C]N); ¹H NMR (DMSO-d₆, d, ppm): 1.66 (t, 2H,

J ¼ 6.02 Hz, CH₂), 2.29 (t, 2H, J ¼ 6.00 Hz, CH₂), 2.65e

2.70 (m, 2H, CH₂), 2.83 (s, 3H, SCH₃), 3.29e3.33 (m, 2H,

CH₂), 3.35e3.38 (m, 4H, CH₂), 3.45e3.49 (m, 2H, CH₂),

7.05 (d, 2H, J ¼ 8.37 Hz, phenyl), 7.28 (d, 2H, J ¼ 8.39 Hz,

phenyl), 7.36 (d, 2H, J ¼ 8.40 Hz, phenyl), 7.53 (d, 2H,

J ¼ 8.41 Hz, phenyl), 8.27 (s, 1H, CH), 9.60 (br, NH, D₂O

exchangeable); its MS, [M^þ], m/z 543 (100%), [M^þþ 2], m/z

545 (22%), [M^þþ 4], m/z 547 (9%).

(CH alkyl), 1708 (CO), 1630 (C]N); H NMR (DMSO-d₆,

d, ppm): 1.69 (t, 2H, J ¼ 6.01 Hz, CH₂), 2.28 (t, 2H, J ¼

6.03 Hz, CH₂), 2.68e2.75 (m, 2H, CH₂), 2.83 (s, 3H,

SCH₃), 6.94e7.15 (m, 4H, phenyl), 7.21 (d, 2H, J ¼

8.41 Hz, phenyl), 7.32e7.38 (m, 2H, phenyl), 7.52 (d, 1H,

J ¼ 9.07 Hz, phenyl), 7.62 (d, 2H, J ¼ 8.38 Hz, phenyl), 7.73

(d, 1H, J ¼ 9.23 Hz, phenyl), 8.45 (s, 1H, CH), 9.50 (br,

NH, D₂O exchangeable); its MS, [M^þ], m/z 580 (100%),

[M^þþ 2], m/z 582 (38%), [M^þþ 4], m/z 584 (15%).

5.1.21. Synthesis of 5-(4-chlorophenyl)-9-

(4-chlorophenylmethylene)-2,4-dihydrazino-6,7,8,9-

tetrahydropyrimido[4,5-b]quinoline (23)

A mixture of 18 (0.01 mol) and hydrazine hydrate (99e

100%, 10 mL) was stirred under reﬂux in dioxane (30 mL)

and ethanol (5 mL) for 12 h. The reaction mixture was allowed

to cool to 0 ^ꢃC for 5 h, and the solid was collected by ﬁltration

and crystallized from dioxane as a pale yellow powder. IR (KBr,

5.1.18.5. Synthesis of 5-(4-chlorophenyl)-9-(chlorophenyl-

methylene)-4-morpholino-2-methylthio-6,7,8,9-tetrahydropyri-

mido[4,5-b]quinoline (20b). It was obtained from morpholine

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

623

cm^ꢀ1): 3438e3350 (br s, NH’s), 3035 (CH aryl), 2923 (CH

alkyl), 1625 (C]N); H NMR (DMSO-d₆, d, ppm); 1.71 (t,

prepared in phosphate buffer (pH 7, 0.1 M). The mixture was

shaken, centrifuged, ﬁltered, and the absorbance (A_control) of

the resulting green-blue solution (ABTS radical solution)

was adjusted at ca. 0.5 at l 734 nm. Then, 50 ml of (2 mM)

solution of the test compound in spectroscopic grade MeOH/

1

2H, J ¼ 5.97 Hz, CH₂), 2.26e2.34 (m, 2H, CH₂), 2.45 (br,

NH₂, D₂O exchangeable), 2.75e2.78 (m, 2H, CH₂), 7.07 (d,

2H, J ¼ 8.45 Hz, phenyl), 7.16 (d, 2H, J ¼ 8.42 Hz, phenyl),

7.30 (d, 2H, J ¼ 8.40 Hz, phenyl), 7.43 (d, 2H, J ¼ 8.39 Hz, phe-

nyl), 8.32 (s, 1H, CH), 8.54e8.80 (br, NH₂, D₂O exchangeable),

10.70, 11.60 (two, br s, 2NH, D₂O exchangeable).

phosphate buffer (1:1) was added. The absorbance (A_test

)

was measured and the reduction in color intensity was ex-

pressed as % inhibition. The % inhibition for each compound

is calculated from the following equation [24]:

5.2. Pharmacological screening

A_controlꢀ A_test

% Inhibition ¼

ꢁ 100

5.2.1. Animals

A_control

Male SpragueeDawley rats (150e200 g) were used in the

study of anti-oxidant activity and the adult females were used in

anti-inﬂammatoryactivitystudy. Bothsexof Swissmiceweighing

25e30 g were used in analgesic activity taking into account inter-

national principle and local regulations concerning the care and

useof laboratory animals [22]. Theanimals hadfree access to stan-

dard commercial diet and water ad libitum and were kept in rooms

maintained at 22 ꢂ 1 ^ꢃC with 12 h lightedark cycle.

Ascorbic acid (vitamin C) was used as standard anti-oxi-

dant (positive control). Blank sample was run without ABTS

and using MeOH/phosphate buffer (1:1) instead of sample.

Negative control sample was run with MeOH/phosphate buffer

(1:1) instead of tested compound.

5.2.3.3. Bleomycin-dependent DNA damage. The assay was

done according to Aeschlach et al. [25] with minor modiﬁca-

tions. The reaction mixture (0.5 mL) contained DNA (0.5 mg/

mL), bleomycin sulfate (0.05 mg/mL), MgCl₂(5 mM), FeCl₃

(50 mM) and samples to be tested at different concentrations.

L-Ascorbic acid was used as a positive control. The mixture

was incubated at 37 ^ꢃC for 1 h. The reaction was terminated

by addition of 0.05 mL EDTA (0.1 M). The color was devel-

oped by adding 0.5 mL thiobarbituric acid (TBA) (1%, w/v)

and 0.5 mL HCl (25%, v/v) followed by heating at 80 ^ꢃC for

10 min. After centrifugation, the extent of DNA damage was

measured by increase in absorbance at 532 nm.

5.2.2. Reagents

DNA (Type 1, calf thymus), bleomycin sulfate, butylated

hydroxyanisole (BHA) and ^L-ascorbic acid were obtained

from Sigma. 2,2⁰-Azo-bis-(2-amidinopropane) dihydrochlor-

ide (AAPH) and 2,2⁰-azino-bis(3-ethyl benzthiazoline-6-

sulfonic acid) (ABTS) were purchased from Wak. All other

chemicals were of the highest quality available.

5.2.3. Anti-oxidant screening

5.2.3.1. Assay for erythrocyte hemolysis. Blood was obtained

from rats by cardiac puncture and collected in heparinized

tubes. Erythrocytes were separated from plasma and the buffy

coat and washed three times with 10 volumes of 0.15 M

NaCl. During the last washing, the erythrocytes were centri-

fuged at 2500 rpm for 10 min to obtain a constantly packed

cell preparation. Erythrocyte hemolysis was mediated by per-

oxyl radicals in this assay system [23]. A 10% suspension of

erythrocytes in pH 7.4 phosphate-buffered saline (PBS) was

added to the same volume of 200 mM 2,2⁰-azobis (2-amidino-

propane) dihydrochloride (AAPH) solution (in PBS) containing

samples to be tested at different concentrations. The reaction

mixture was shaken gently while being incubated at 37 ^ꢃC for

wh. The reaction mixture was then removed, diluted with eight

volumes of PBS and centrifuged at 2500 rpm for 10 min. The

absorbance A of the supernatant was read at 540 nm. Similarly,

the reaction mixture was treated with eight volumes of distilled

water to achieve complete hemolysis, and the absorbance B of

the supernatant obtained after centrifugation was measured at

540 nm. The percentage hemolysis was calculated by equation

(1 ꢀ A/B) ꢁ 100%. The data were expressed as mean standard

deviation. ^L-Ascrobic was used as a positive control.

5.2.4. Anti-inﬂammatory activity (carrageenan-induced

rat hind paw edema model)

The method adopted resembles essentially that described

by Winter et al. [26]; distilled water was selected as vehicle

to suspend the standard drugs and the test compounds. The

albino rats weighing between 150 and 180 g was starved for

18 h prior to the experiment. The animals were weighed,

marked for identiﬁcation and divided into 17 groups each

group containing six animals. Edema was induced in the left

hind paw of all rats by subcutaneous injection of 0.1 mL of

1% (w/v) carrageenan in distilled water into their footpads.

The 1st group was kept as control and was given the respective

volume of the solvent (0.5 mL distilled water). The 2nd to

16th groups were orally administered aqueous suspension of

the synthesized compounds in dose of 20 mg/kg (1 h) before

carrageenan injection. The last group (standard) was adminis-

tered diclofenac sodium in a dose of 20 mg/kg orally as aque-

ous suspension [27]. The paw volume of each rat was

measured immediately by mercury plethysmometer, before

carrageenan injection and then hourly for 4 h post-administra-

tion of aqueous suspension of the synthesized compounds. The

edema rate and inhibition rate of each group were calculated

as follows: edema rate (E )% ¼ V_tꢀ V₀/V₀, inhibition rate

(I )% ¼ E_cꢀ E_t/E_c, where V₀is the volume before carrageenan

injection (mL), V_tis the volume at t hours after carrageenan

5.2.3.2. Anti-oxidant activity screening assay e ABTS method.

For each of the investigated compounds 2 mL of ABTS solu-

tion (60 mM) was added to 3 M MnO₂solution (25 mg/mL) all

624

A.B.A. El-Gazzar et al. / European Journal of Medicinal Chemistry 44 (2009) 609e624

[4] Osselaere JP. J. Pharm. Belg. 1974;29(2):145.

injection (mL), E_c, E_tare the edema rate of control group and

treated group, respectively.

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Kohlhaas KL, Wismer CT, Mikusa J, Jarvis MF, Kowaluk EA,

Bhagwat SS. J. Med. Chem. 2003;46:5249.

5.2.5. Analgesic activity using hot-plate test

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The experiment was carried out as described by Turner [28]

using hot-plate apparatus maintained at 53 ꢂ 0.5 ^ꢃC. The mice

were divided into 17 groups of six animals each. The reaction

time of the mice to the thermal stimulus was the time interval

between placing the animal in the hot plate and when it licked

its hind paw or jumped. Reaction time was measured prior to

aqueous suspension of synthesized compounds and drug

treatment (0 min). Group 1 was kept as normal control. The

aqueous suspension of synthesized compounds was orally

administered to mice of groups 2e16 at doses of 20 mg/kg.

Mice of group 17 (reference) were orally treated with diclofe-

nac sodium in a dose of 20 mg/kg body weight. The reaction

time was again measured at 15 min and repeated at 30, 60 and

90 min after treatment. To avoid tissue damage to the mice

paws, cut-off time for the response to the thermal stimulus

was set at 60 s. The reaction time was calculated for each

synthesized compound and drug-treated group.

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writhing response model)

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The compounds were selected for investigating their analge-

sic activity in acetic acid induced writhing response in Swiss

albino mice following the method of Collier et al. [29]. One hun-

dred and two mice were divided into 17 groups (six in each

group), starved for 16 h and pretreated as follows: the 1st group

which served as control positive was orally received distilled

water in appropriate volumes. The 2nd to 16th groups were

received the aqueous suspension of synthesized compounds

orally in a dose of 20 mg/kg. The last group was orally received

diclofenac sodium in a dose of 20 mg/kg. After 30 min, each

mouse was administered 0.7% of an aqueous solution of acetic

acid (10 mL/kg) and the mice were then placed in transparent

boxes for observation. The number of writhes was counted for

20 min after acetic acid injection. The number of writhes in

each treated group was compared to that of a control group.

The number of writhing was recorded and the percentage protec-

tion was calculated using the following ratio: % protection ¼

(control mean ꢀ treated mean/control mean) ꢁ 100.

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Article Doi

Design and synthesis of azolopyrimidoquinolines, pyrimidoquinazolines as anti-oxidant, anti-inflammatory and analgesic activities

DOI: 10.1016/j.ejmech.2008.03.022

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Full text of DOI:10.1016/j.ejmech.2008.03.022

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