SYNTHESIS OF THE N1-SUBSTITUTED QUINOLONES, N1

QUINOLONE DIMERS AND C-6 SUBSTITUTED PYRAZOLO

PYRIMIDINES AS ANTIBACTERIAL AGENTS

A SYNOPSIS

Submitted

in partial fulfillment of the requirement for the award of the degree of

DOCTOR OF PHILOSOPHY

in

FACULTY OF CHEMISTRY

By

C. NAVEENKUMAR REDDY

[Reg. No.410410 C/PH]

RESEARCH AND DEVELOPMENT CELL

JAWAHARLAL NEHRU TECHNOLOGICAL UNIVERSITYHYDERABAD

KUKATPALLY, HYDERABAD – 500 085

INDIA

JUNE 2011

Synopsis

1

SYNOPSIS

of the

Ph. D Thesis

entitled

SYNTHESIS OF THE N1-SUBSTITUTED QUINOLONES, N1

QUINOLONE DIMERS AND C-6 SUBSTITUTED PYRAZOLO

PYRIMIDINES AS ANTIBACTERIAL AGENTS

Biologically active compounds are isolated either from natural products or

prepared by chemical synthesis. These active compounds are widely used

for survival of living things. These compounds are classified as agonists,

antagonists, antibacterials, cardiovascular, antifungals and anticancers.

Among all the classes, antibacterial agents are largest and most

important therapeutic area, which cures infectious diseases. Prolonged

usage of these antibacterial agents leads to the resistance in the bacterial

species. Hence, continuous development is needed in this area.

The high importance for the novel antibacterial agents needed to society,

motivated us to work in this area of interest. In this thesis we wish to

report introduction on evolution of antibacterial agents (Chapter 1),

synthesis of some novel potential antibacterial agents, such as N-

substituted quinoline-3-carboxylic acids and corresponding amides

(Chapter 2), N-1-dimers of quinoline-3-carboxylic acids (Chapter 3),

Quinazolino-quinolones (Chapter 4) and Pyrazolopyrimidine derivatives

(Chapter 5). Also we have synthesized various quinolone-3-carboxamides

(Chapter 2) which are known to have activity towards cannabinoid

receptor agonist.

Synopsis

2

1.0 CHAPTER 1: INTRODUCTION TO ANTIBACTERIAL AGENTS

In this chapter we have described about the identification and

classification of bacteria and mode of action of antibacterial agents. We

have also described about the evolution of antibacterial agents from sulfa

drugs, penicillins, cephalosporins, tetracyclines to quinolones. The

literature reports available for structure activity relationships (SAR) of

quinolone derivatives have been demonstrated. The recent reports on

pyrazolopyrimidnes as new class of antibacterials were cited in this

chapter.

2.0 CHAPTER 2: SYNTHESIS OF NOVEL N-SUBSTITUTED-1,4-

DIHYDRO-4-OXO-3-QUINOLINECARBOXYLIC ACIDS AND

AMIDES

Since the discovery of the quinolinone anti-bacterial agent nalidixic acid

in 1962, a large number of its congeners have been synthesized and

extensively evaluated for their structure-activity relationship studies,

which led to a number of new analogues with dramatic improvement of

their biological activity. Such modifications have been a subject of current

interest to synthetic and medicinal chemists.

N-alkyl/aryl-1,4-dihydro-4-oxo-3-quinoline carboxylic acids and its

ester derivatives are key building blocks of quinolinone anti-bacterial

agents, which are effective against a variety of microorganisms, including

gram-negative and gram-positive pathogens. The recent interesting

results reported on analogues of quinolone class of carboxamides

biological activity on HIV-1, NiV, HSV-1, HSV-2 and cannabinoid CB2

Synopsis

3

receptor agonist are aggravated us to synthesize a novel N-substituted-

1,4-dihydro-4-oxo-3-quinoline carboxylic acid and amide derivatives

2.1 Synthesis of novel N-substituted-1,4-dihydroquinoline

carboxylic acids

Herein, we present an expeditious synthesis of novel N-substituted-1,4-

dihydro-4-oxo-3-quinoline carboxylic acids in five steps from a

commercially available 2-bromobenzoic acid via regiospecific cyclisation of

ethyl-3-(2-bromophenyl)-2-((N-substituted)methylene)-3-oxo propanoate.

2.1.1 Synthesis of ethyl-3-(2-bromophenyl)-2-((dimethylamino)

methylene)-3-oxopropanoate

2-bromo benzoic acid (2.1) was converted into corresponding acid

chloride (2.2) in the presence of thionyl chloride followed by condensation

with potassium salt of monoethyl malonate (2.3) to obtain bromobenzoyl

β- keto ester (2.4) which was further reacted with N,N-dimethylformamide

dimethyl acetal to give ethyl-3-(2-bromophenyl)-2-((dimethylamino)

methylene)-3-oxopropanoate (2.6) in about 33% over all yield as shown in

scheme-2.1

In an alternate process to minimize number of steps ethyl 3-

(dimethylamino) acrylate was introduced to build quinolinone skeleton.

Bromobenzoyl chloride (2.2) was reacted with ethyl-3-(dimethyl

amino)acrylate (2.5) in the presence of triethylamine and acetonitrile to

give ethyl-3-(2-bromophenyl)-2-((dimethylamino) methylene)-3-

oxopropanoate (2.6) in about 48% over all yield.

Synopsis

4

Br

OH

OO

O

O

Br

O

O

O

Br N

O O

OKO

Br

O

Cl

O

O N

2.1 2.2

2.3

2.4

2.6

Thionylchloride/ DMF/ DCM

RT 0-5°C

MgCl2/ TEA/ ACN

DMF/ DMA

50-55°C

TEA/ ACN

0°C - RT

2.5

Scheme 2.1 Synthesis of ethyl-3-(2-bromophenyl)-2-((dimethylamino)

methylene)-3-oxopropanoate.

2.1.2 Synthesis of ethyl-4-oxo-N-pyridyl-1, 4-dihydroquinoline-3-

carboxylate

Condensation of ethyl-3-(2-bromophenyl)-2-(dimethylamino) methylene)-

3-oxopropanoate (2.6) with 2-aminopyridine, 3-aminopyridine and 4-

aminopyridine undergoes insitu cyclization to afford ethyl-4-oxo-N-

pyridyl-1,4-dihydroquinoline-3-carboxylate (2.7-2.9) derivatives as

shown in scheme 2.2.

N

O O

O

R

O

Br

O

O

N

N

NN

2.6

Reflux Y: 55%

2.7 R =

Aminopyridines/Cesium carbonate/ TEA/acetonitrile

2.8 R = 2.9 R =

Scheme-2.2: Synthesis of ethyl-4-oxo-N-pyridyl-1, 4-dihydroquinoline -3-

carboxylate (2.7-2.9)

Synopsis

5

2.1.3 Synthesis of ethyl-4-oxo-N-substituted-1,4-dihydro quinoline-3-

carboxylate

Ethyl-3-(2-bromophenyl)-2-((dimethylamino)methylene)-3-oxopropanoate

(2.6) on condensation with 4-phenylbutan-2-amine, 4-

(aminomethyl)pyridine, 2-aminothiazole and 1-amino-4-methyl piperazine

obtained an intermediate ethyl-3-(2-bromophenyl)-2-(( N-

alkylamino)methylene)-3-oxo propanoate (2.10-2.13). The open forms

were then converted to quinolinone ester (2.14 and 2.15) by further

treatment with cesium carbonate and catalytic amount of copper iodide as

shown scheme 2.3.

N

O O

O

R

O

Br

O

O

N

O

Br

O

O

N

R

N

CH3 CH

3

S

N

NN CH3

N

2.62.10 R=

2.11 R=

2.14 R=

2.15 R=

Amine/ TEA

Reflux Y: 70-80%

TEA or Cs2CO3 or

Cs2CO3 / CuI

Reflux Y: 65-75%

2.12 R=

2.13 R=

Scheme-2.3 Synthesis of ethyl-4-oxo-N-substituted-1,4-dihydro

quinoline-3-carboxylate

2.1.4 Synthesis of 4-oxo-N-substituted-1,4-dihydroquinoline-3-

carboxylic acids

Compounds (2.7-2.9 and 2.14-2.15) were hydrolyzed in presence of

aqueous NaOH to obtain respective N-alkyl/ary-1,4-dihydro-4-oxo-3-

Synopsis

6

quinolinecarboxylic acids (2.16-2.20) in about 55-75 % as shown in

scheme 2.4.

N

O O

O

RN

O O

OH

R

N

N

N

N

N

N

N

N

20% NaOH solution

2.14 R=

2.15 R=

2.7 R=

2.8 R=

2.9 R=

2.19 R=

2.20 R=

2.16 R=

2.17 R=

2.18 R=

RefluxY: 70-80%

Scheme 2.4 Synthesis of 4-oxo-N-substituted-1,4-dihydroquinoline-3-

carboxylic acids

2.2 Novel N-substituted-1,4-dihydroquinoline carboxamides as

potential anti viral and CB2 Cannabinoid receptor agonists

Quinolone class of carboxamides has been shown to inhibit HIV-1.

Interestingly, this quinolone class of compounds presents an extremely

low cytotoxic effect compared to other class. Recent reports on molecular

modeling and pharmacological characterization studies reveal that 4-oxo-

1, 4-dihydro quinoline-3-carboxamide derivatives are active as potent

CB2-selective receptor ligands. A general and high yielding synthetic route

for the preparation of novel N-pyridyl- 1, 4-dihydro-4-oxo-3-

quinolinecarboxamide derivatives (2.21-2.33) have been demonstrated in

scheme-2.5.

Synopsis

7

N

O O

OH

R

NH

N

O O

R

R1

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

CH3

N

CH3

CH3

N

CH3

N

CH3

N

CH3

N

CH3

N

CH3

CH3

N

CH3

N

R1-Amine/HATU/diisopropylethyl amine/ DMF/DCM

R1= 2.29 R=

2.31 R=

2.21 R=

2.23 R=

2.26 R=

2.19 R=

2.20 R=

2.16 R=

2.17 R=

2.18 R=

2.22 R=

2.24 R=

2.25 R=

2.27 R=

2.28 R=

2.30 R=

2.32 R=

2.33 R=

R1=

R1=

R1=

R1=

R1=

R1=

R1=

R1=

R1=

R1=

R1=

R1=

Scheme 2.5 Synthesis of 4-oxo-N-sustituted-1,4-dihydroquinoline-3-

carboxamides

The synthesis of N-substituted-1,4-dihydro-4-oxo-3-quinoline carboxylic

acids (2.16-2.20) were reported in the section 2.1.4 of this chapter. These

carboxylic acids were coupled with variety of amines such as 4-

(aminomethyl) pyridine (primary amines), phenylethylamine (secondary

amine) and 2-methyl-aniline (aromatic amine) to afford 4-oxo-N-

sustituted-1,4-dihydroquinoline-3-carboxamides (2.21-2.33). The

coupling agent used in this process was HATU, high purity products were

obtained in good yields ranging from 50-75%. The reaction yields have

been improved when we have changed the base from TEA to DIPEA.

2.3 Conclusion

In conclusion, a novel N-sustituted-1,4-dihydro-4-oxo-3-quinoline

carboxylic acids and corresponding amides were synthesized with an

Synopsis

8

efficient synthetic route. The purpose of this investigation was to evaluate

new and high potential quinolone class of anti bacterial and CB1

cannabinoid receptor agonists.

3.0 CHAPTER 3: QUINOLONE DIMERS AS POTENTIAL ANTI

BACTERIAL AGENTS

The importance of quinoline class of anti bacterials motivated us to

choose as area of interest towards synthesis of N1-quinolone dimers

(fig.3.1) with two carbon chain linker. Pharmacokinetic studies indicate

that the bulky substitution at N1 position will enhance the bioavailability

of the quinoline-3-carboxylic acids.

Figure 3.1

N N CH3NHN NHN

NH

N

ONa

O

N

O

ONa

O

F

F

N

O

F

F

R

R

R=

As number of quinolone anti-bacterials contains 1, 4-dihydro-4-oxo-3-

quinolinecaroboxylic acid as a basic scaffold, in general most of the

modifications were carried on C-7 of quinolone and N1 of the quinolone to

enhance their activity. Keeping the substitution at N1-position having a

STERIMOL length of 0.42 nm, is an important parameter for better

activity. The linkers such as 1, 2-diaminoethane and 1, 2-

diaminocyclohexane were used to prepare the dimers.

Synopsis

9

3.1 Synthesis of fluoro quinolone dimers

An exploratory synthesis of N1-dimer analogues of 6-fluoro-1, 4-dihydro-

4-oxo-3-quinolinecarboxylic acid derivatives from corresponding benzoic

acids via corresponding acrylates were demonstrated successfully with

good overall yields. Both the methods (where R = H) for the synthesis of

corresponding acrylates (3.9-3.11) have been used to make N1-dimers to

compare the overall yields and it has been observed that starting from

acetophenone method is better than starting from benzoic acid (Scheme

3.1) method.

F

OH

O

F

F

R

O

O

O

F NF

F

R

F

O

Cl

F

F

R

F

O

F

FO

O

O

FF

F

3.3 R = H3.4 R = F3.5 R = OCH3

3.6 R = H 3.7 R = F3.8 R = OCH3

3.9 R = H3.10 R = F3.11 R = OCH3

SOCl2/DMF/

Toluene

55-60°CY: 75-92%

Ethyl-3-(dimethyl amino)acrylate/TEA/ Toluene

55-60°C Y: 30-48%

3.1

NaH/(C2H50)2CO

Y: 90%DMF-DMA

50-55°C, 2 h, Y: 91%3.2

Scheme 3.1. Synthesis of acrylates

3.2 Synthesis of ethyl linker fluoro quinolone N1-dimers

Acrylates 3.9, 3.10 and 3.11 were treated with half equivalent of 1,2

diamino ethane in acetonitrile at ambient temperature results in

formation of dimeric enamine ketoesters 3.14, 3.15 and 3.16. In these

reactions the ratio of the ethylene diamine (0.5 eq and 1 eq.) plays an

important role in resulting different products. It has been demonstrated

at first the regiospecific dimeric cyclisation to form favored 6-membered

Synopsis

10

ring resulted in N1-dimers of 6,7-difluoro-1,4-dihydro-4-oxo-3-quinoline

carboxylates 3.17, 3.18 and 3.19 (Scheme 3.2). N1-dimers 3.17, 3.18

and 3.19 were hydrolyzed using aqueous sodium hydroxide in THF at

reflux temperature to produce the corresponding diacids 3.20, 3.21 and

3.22.

O

O

O

F N

R

F

F

O O

O

R

F

F

NH

O

O

O

R

F

F

NH

F

F

O

N

O

O

O

R

F

F

N

O

O

R

F

F

O

N

O

OH

O

R

F

F

N

O

OH

R

F

F

3.14 R=H3.15 R=F3.16 R=OCH3

3.20 R=H3.21 R=F3.22 R=OCH3

3.17 R=H3.18 R=F3.19 R=OCH3

3.9 R=H3.10 R=F3.11 R=OCH3

Ethylenediamine/ACN

RT, 3 hY: 50-65%

Cs2CO3/ACN

20% NaOH sol./THF

Reflux, 4 hY: 53-66%

Reflux, 4 hY: 80-85%

Scheme 3.2. Synthesis of ethyl linker fluoro quinolone N1-dimers

3.3 Synthesis of ethyl linker 6-fluoro-7-substituted-1,4-dihydro-4-

oxo-3-quinoline carboxylic acid N1-dimers (3.26-3.29)

For further enhancement of the activity of fluoroquinolone dimers,

fluorine at C-7 position was replaced with various amines as shown in

Scheme 3.3. Displacement of C-7 fluorine on 3.17 with an excess

Synopsis

11

amount of amines at 80-85°C result compounds 3.23-3.25. However, we

could not isolate and purify these compounds due to its poor solubility in

common organic solvents.

O

N

O

O

O

H

F

F

N

O

O

H

F

FO

N

O

OH

O

H

F

F

N

O

OH

H

F

F

O

N

O

OH

O

H

F

N

O

OH

H

F

R1

R1

N N

NHN

O

O

N

O

O

O

H

F

N

O

F

H

R1

R1

NN

N

N N

NHN

NN

Amine Amine

3.20 3.17

3.27 R1=

3.26 R1=

N-methylpyrrolidinone

80-85°C, 5hY: 45-68%

3.28 R1=

Aq. NaOH/THF

N-methylpyrrolidinone

80-85°C, 5h

3.29 R1=

3.24 R1=

3.23 R1=

3.25 R1=

Reflux, 4 hY: 56%

X

Scheme-3.3. Synthesis of ethyl linker fluoro quinolone N1-dimers

We have hydrolyzed the compound 3.17 in to 6,7-difluoro-1,4-dihydro-4-

oxo-3-quinolinecarboxylicacid 3.20. The C-7 position of compound 3.20

was substituted with amines at 800C for 4 h to afford compounds 3.26,

3.27, 3.28 and 3.29 which were isolated as free solids, also with poor

solubility in common organic solvents.

Synopsis

12

3.4 Synthesis of ethyl linker 6, 8-difluoro-7-substituted amino-1,4-

dihydro-4-oxo-3-quinoline carboxylic acid N1-dimers and its

derivatives (3.34-3.37).

Displacement of C-7 fluorine on 3.18 with an excess amount of amines at

80-85°C yielded respective C-7 substituted dimers 3.30-3.33 in good

yields, followed by hydrolysis with 20% NaOH in methanol gave

corresponding acids 3.34 – 3.37

O

N

O

O

O

F

F

F

N

O

O

F

F

F

O

N

O

OH

O

F

F

N

O

OH

F

F

R1

R1

N N CH3

NHN NHN

NH

N

O

O

N

O

O

O

F

F

N

O

F

F

R1

R1

N N CH3

NHN NHN

NH

N

3.18 3.30 R1=

3.31 R1=

3.32 R1=

3.33 R1=

Amine/ N-methylpyrrolidinone

80-85°C, 5hY: 55-60%

20% NaOH sol./MeOH

Reflux, 5 hY: 54-90%

3.34 R1=

3.35 R1=

3.36 R1=

3.37 R1=

Scheme-3.4. Synthesis of ethyl linker fluoro quinolone N1-dimers

3.5 Synthesis of cyclohexyl linker fluoro quinolone N1-dimers

Synopsis

13

O O

O

F

F

N

N

O

O

O

F

F

F

F

R

R

O

O

O

F NF

F

R

O

O

N

O

O

O

F

F

N

R

F

F

R

O

OH

O

N

O

OR'

O

F

F

N

R

F

F

R

O

3.10 R=F3.11 R=OCH3

Cyclohexane-1,2-diamine/Cs2CO3/ACN

Reflux, 30 minY: 39-50%

6N HCl sol./ethanolor 20% aq. NaOH sol.THF

Reflux, 2h Y: 55%

3.38 R=F3.39 R=OCH3

3.40 R=F3.41 R=OCH3

3.42 R=F R'=C2H5

3.43 R=OCH3 R'= H

Scheme-3.4. Synthesis of cyclohexyl linker fluoro quinolone N1-dimers

Compounds 3.10 and 3.11 (Scheme 3.4) were treated with half

equivalent of cyclohexyl-1, 2-diamine in acetonitrile, which results

compounds 3.40 and 3.41. Compound 3.40 was treated with aq. HCl or

NaOH in ethanol at reflux resulted in mono-acid 3.42 in 44% isolated

yield and rest was unreacted starting material, and it was observed that

this was not converting in to di acid. The increase in the basicity or

temperature of the reaction is leading to decomposition.

Antibacterial activity of fluoro quinolone dimers

Compounds (3.17-3.19) were insoluble in test solution, hence activity test

was not performed, and interestingly compounds 3.22 and 3.43 does not

Synopsis

14

show activity towards Gram negative and does show activity over Gram

positive. Compounds 3.20, 3.26-3.33 and 3.42 do show activity towards

Gram negative and does not show activity over Gram positive.

Compounds 3.40, 3.41 were shown zero activity for both the strains. All

other compounds (3.21, 3.34-3.37) were shown approximately half of the

activity when compared with standard (Ciprofloxacin).

Conclusion

Dimer analogs of 1,4-dihydro-4-oxo-7-substituted amino-6-

fluoroquinoline-3-carboxylic acids and its ester derivatives are effective

against a variety of gram negative microorganisms. The in-vitro

antibacterial activity result shows that these dimers are having moderate

activity when compared with Ciprofloxacin.

4.0 CHAPTER 4: SYNTHESIS OF NOVEL QUINAZOLINO-

QUINOLONES AS POTENCIAL ANTIBACTERIAL AGENTS

In view of wide veriety of pharmaceutical applications with quinazolinone,

we have designed and synthesized a novel series of quinazolinono-

quinolone derivatives (fig 4.1) to study the antibacterial activity as well as

other therapeutic areas.

Figure 4.1

O

O

O

F

NF

R1

N

N

O

R

R1= H, OCH3

R = H, Cl

Synopsis

15

Here, we wish to report the synthesis of 3-quinazolinones coupled with N-

1 position of 6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylate with a

two carbon chain linker to evaluate the biological activities. Interestingly

these compounds have two core components viz. quinazolinone (core

component 1) and quinolone (core component 2)

4.1 Synthesis of N-ethylamino quinazolinone core component

NH2

O

OHR

N

O

O

R

N

N

O

R NH2

N

N

O

N

N

O

R R

4.1 R= H4.2 R= Cl

4.3 R=H4.4 R=Cl

Acetic anhydride

110-140°C, Y: 76-78%

1,2 diaminoethane

ACN/K2CO3

X

1,2 diaminoethane

ACN/K2CO3

4.6 R=H

4.5 R=H

Scheme-4.1. Synthesis of quinazolinone core component

Anthranilic acids (4.1 and 4.2) were treated with acetic anhydride at 110-

140°C to afford lactones (4.3 and 4.4), in quantitative yields. The

obtained lactones were treated with 1 eq of 1,2-diaminoethane resulted in

formation of quinazolinone dimer (4.6) instead of N-ethylamino

quinazolinone (4.5). The same dimer was obtained even though we have

increased the quantity of 1,2-diaminoethane. As the above reaction

resulting in formation of 4.6 instead of 4.5, an alternate route was

proposed for the synthesis of quinazolino-quinolones as shown below.

Synopsis

16

4.2 Synthesis of N-ethyl amino quinolones

N-ethyl amino quinolones (core component 2) were prepared from a key

intermediate acrylates (3.9 and 3.11) which were synthesized in chapter

3 from corresponding fluorobenzoic acids. Compound 3.9 and 3.11 were

reacted with one equivalent of 1,2 diamino ethane in acetonitrile at

ambient temperature resulted in formation of corresponding enamine

ketoesters 4.7 and 4.8. When we used 0.5 equivalent of 1,2-

diaminoethane we got exclusively dimers as in chapter 3. Regiospecific

cyclisation resulted in N-ethylamino quinolones 4.9 and 4.10.

O

O

O

F NF

F

R

O

O

O

F

F

R

N

NH2

F

O

O

O

F

F

R

N

NH2

3.9 R=H3.11 R=OMe

Ethylenediamine

Acetonitrile

RT

Cesium carbonate

80°C-85°C, Y: 70-90%

4.7 R=H4.8 R=OMe

4.9 R=H4.10 R=OMe

Scheme-4.2. Synthesis of N-ethyl amino quinolones

4.3 Synthesis of quinazolino-quinolone

Compound 4.9 and compound 4.10 were treated with lactones 4.3 and

4.4 in ethanol resulted in corresponding products 4.11 to 4.14 in

moderate to good yields. The esters 4.11 to 4.14 were tried to hydrolyze

using basic and acidic media and found that these compounds are

decomposing to multiple spots and could not isolate the corresponding

acids (4.15 to 4.18).

Synopsis

17

O

O

O

N

R1

F

F

NH2

N

N

O

O

O

R1

F

F

N

O

R

N

O

O

R

N

N

O

O

O

F

F

N

O

R1

R

4.11 R=H, R1=H4.12 R=Cl, R1=H4.13 R=H, R1=OCH3

4.14 R=Cl, R1=OCH3

+

4.3 R=H4.4 R=Cl

Ethanol

Reflux, Y: 16-27%

Aq. NaOH

X

4.9 R=H4.10 R=OCH3

4.15 R=H, R1=H4.16 R=Cl, R1=H4.17 R=H, R1=OCH3

4.18 R=Cl, R1=OCH3

Scheme-4.3 Synthesis of quinazolino-quinolones

Antibacterial activity of quinazolino- quinolone

All compounds (4.11-4.14) were shown approximately half of the anti

bacterial activity when compared with standard (Ciprofloxacin).

4.4 Conclusion

Quinazolinone derivatives are well known anti microbial agents such as

anti bacterial, anti fungal, anti malarial and anti cancer agents. In view of

high importance of these analogues we have synthesized a new series of

quinazolino-quinolones with a two carbon chain linker to increase their

anti bacterial activity.

Synopsis

18

5.0 CHAPTER 5: NOVEL PYRAZOLOPYRIMIDINES AS POTENTIAL

ANTIBACTERIAL AGENTS

Various pyrazolopyrimidines reported are active towards microbial agents

and antibacterial activity. There are very few methods for the synthesis of

monochloropyrazolopyrimidines in the literature. We have developed a

new pathway for the synthesis of substituted pyrazolopyrimidines (fig 5.1)

from barbituric acid.

Figure-5.1

N

N N

N

RX

R1

R3

R2

R4

R = H, C6H5

R3 = H, Cl, F, OCH3

R2 = H, CH3, F,

R1 = H, NH2

R4 = H, CH3, Cl

X = O, NH

5.1 Synthesis of mono chloro pyrazolopyrimidine and its

derivatives

Barbituric acid (5.1) was treated with POCl3 in DMF at 80-85°C to obtain

5-formyl-2,4,6 trichloropyrimidine (5.2) with 60% yeild. The aldehyde 5.2

was further treated with corresponding hydrazine hydrate in presence of

base such as triethylamine, resulted is dichloropyrazolopyrimidine (5.3

and 5.4) around 71% yield. The obtained dichloropyrazolopyrimidine 5.3

was de-chlorinated selectively at the C-4 position using H2-Pd/C and

sodium triacetoxy borohydride in ethanol and compound 5.4 was

converted using H2-Pd/C and triethylamine at 60 psi for 4 hrs to obtain

Synopsis

19

corresponding monocholropyrazolopyrimidine (5.5 and 5.6) in about 30%

isolated yield as shown in scheme 5.1.

N

NOH

OH

OH

N

NCl

Cl

Cl

H

O

N

NCl

Cl

NH

N

N

NCl

H

NH

N

N

NCl NN

Cl

N

NCl NN

5.1 5.2 5.3

5.55.45.6

-25 to -20°C, Y: 71%

Hydrazine hydratePOCl3/DMF

80-85°C, Y: 60%

H2-Pd/C, TEA

EtOH

H2-Pd/C,

Na(OAc)3BH

EtOH, RTY; 30%

RT, Y: 36%

Phenyl hydrazine

-25 to -20°C,Y: 64%

Scheme-5.1 Synthesis of monochloropyrimidines from barbituric acid

Compound 5.2 was treated with methyl hydrazine and 1(6-Bromo-2-

pyridyl)hydrazine and interestingly we have isolated unexpected

compounds (5.7 and 5.8) as shown below with 80% and 34% isolated

yields respectively even at low temperatures such as -60°C.

N

NCl

Cl

Cl

H

O

N

NCl

NH

NN

CH3

NH

CH3

N

NCl NN

N

NHNH

N

Br

Br

Methylhydrazine,TEA

Methanol

-25 to -20°C, Y: 34%

1(6-Bromo -2-pyridyl)hydrazine,TEA, Methanol

5.75.85.2

-25 to -20°C, Y: 80%

Scheme-5.2 Synthesis of monochloropyrazolopyrimidines derivatives

Synopsis

20

5.2 Alternative route for the synthesis of monochloro

pyrazolopyrimidine

We tried an alternate route for the synthesis of

monochloropyrazolopyrimidines by substituting the 4-chloro functional

group in to 4-amino functional group followed by de-amination using

diazotization method in the presence of copper oxide, which was resulted

in multiple spots. TLC showed slight product but we could not isolate the

desired product 6-chloro-1N-substituted pyrazolopyrimidines (5.5 and

5.6).

N

NCl

Cl

NN

R

N

NCl

NH2

NN

R

N

NCl NN

R

5.3 R=H5.4 R=C6H5

5.9 R=H5.10 R=C6H5

5.5 R=H5.6 R=C6H5

Aq. NH4OH soln

50°C, Y 34%

CopperoxideOrthophosphoric acid

Scheme-5.3 Alternative route for the synthesis of monochloro

pyrazolopyrimidines

5.3 Synthesis of C-6 substituted pyrazolopyrimidine analogs as

potential anti bacterial agents

Monochloropyrazolopyrimidines (5.5 and 5.6) were treated with different

substituted anilines (5.11 – 5.15) to replace chlorine at C-6 carbon to

afford substituted pyrazolopyrimidines (5.16 -5.25).

Synopsis

21

N

NCl NN

R

N

NN NN

R

N

NN NN

MeOR

N

NN NN

F

MeO

R

N

NN NN

FR

N

NN NN

Cl

F

R

NH2

F

NH2

NH2

MeO

NH2

F

MeO

NH2

Cl

F

NaHCO3,DMF

100°C

NaHCO3,DMF,

NaHCO3or TEA,DMF,100°C

Xantphos,Pd(OAc)2,KOtBu

1,4-Dioxane

1,4-Dioxane

Xantphos, Pd(OAc)2, KOtBu

100°C

100°C

100°C

5.11

5.12

5.135.14

5.15

5.16 R= H, 5.17 R=C6H5

5.18 R= H, 5.19 R=C6H5

5.20 R= H, 5.21 R=C6H55.22 R= H, 5.23 R=C6H5

5.24 R= H, 5.25 R=C6H5

5.5 R= H, 5.6 R=C6H5

Scheme-5.4 Synthesis of C-6 substituted pyrazolopyrimidines

5.4 Synthesis of C-4 and C-6 substituted pyrazolopyrimidine

analogs as potential anti bacterial agents

4-amino-6-chloro-pyrazolopyrimidines (5.10) were treated with different

phenols (5.26 - 5.28) to substitute chlorine at C-6 carbon of

pyrazolopyrimidine to obtain new series 4-amino-6-aryloxy-

Synopsis

22

pyrazolopyrimidines (5.29 – 5.31) in moderate yields as shown below.

N

NCl NN

NH2

N

NO NN

N

CF3

NH2

N

NO NN

F

NH2

N

NO NN

F

NH2

Cl

OH

F

OH

F

Cl

OH

N

CF3

Cs2CO3, DMF

100°C

Cs2CO3, DMFCs2CO3, DMF

100°C

5.28 5.26

5.27

5.31 5.29

5.30

100°C

5.10

Scheme-5.5 Synthesis of C-6 substituted 4-amino-1N-phenyl

pyrazolopyrimidines

Antibacterial activity of C-6 substituted pyrazolopyrimidines

Compounds 5.29 showed zero activity towards Gram positive

interestingly does show the activity towards Gram negative. All other

compounds (5.19, 5.20, 5.24, 5.25, 5.31) were shown approximately

half of the activity when compared with standard (Ciprofloxacin).

5.5 Conclusion

The literature reports reveal that pyrazolopyrimidines also show

antibacterial activity. The high demand for the novel antibacterials

motivated us to synthesize these new series of pyrazolopyrimidines.