ABSTRACT

The thesis entitled “Studies on the synthesis of pharmacophoric heterocyclic hybrids

as potent inhibitors of Angiotensin Converting Enzyme (ACE) and Mycobacterium

tuberculosis.” has been divided into six chapters

Chapter I : Introduction to pharmacophoric heterocyclic hybrids: Application

in medicinal chemistry.

Chapter II : Synthesis and evaluation of 2-butyl-4-chloro-1-imidazole

embedded chalcones and pyrazole as Angiotensin Converting

Enzyme (ACE) inhibitors.

Chapter III : Synthesis and evaluation of novel 2-butyl-4-chloro-imidazole

derived peptidomimics as Angiotensin Converting Enzyme (ACE)

inhibitors.

Chapter IV : Synthesis and evaluation of novel 2-hydroxypyrrolobenzo-

diazepine-5,11-dione analogues as potent Angiotensin Converting

Enzyme (ACE) inhibitors.

Chapter V : This chapter subdivided into two sections.

Section A : Synthesis of dihydrobenzofuran tethered pyridines and dihydro-6H-

quinolin-5-ones from β-enaminones as potent inhibitors of

Mycobacterium tuberculosis.

Section B : Synthesis of 6-substituted 2-picolines from aryl / heteroaryl -β-

enaminones and meldrum’s acid.

Chapter VI : This chapter subdivided into two sections.

Section A : Synthesis and antimicrobial evaluation of novel 1-benzyl 2-butyl-4-

chloro imidazole embodied 4-Azafluorenones.

Section B : Synthesis of novel 2-(trifluoromethyl)phenothiazine-[1,2,3]

triazoles and [1,2,3]-triazole1-Adamantylacetamide hybrids as

potent inhibitors of Mycobacterium tuberculosis.

Abstract

i

Chapter I : Introduction to pharmacophoric heterocyclic hybrids:

Applications in medicinal chemistry

Introduction

Drugs were discovered through identifying the active pharmacophoric unit from

traditional remedies and by serendipitous discovery. Modern drug discovery involves the

identification of screening hits and optimization of those hits to increase affinity,

selectivity efficacy/potency, metabolic stability and oral bioavailability. A current trend in

the development of new clinically effective drug candidates is through molecular

modification of known and existing lead nucleus.

Molecular hybridization

Molecular hybridization is a structural modification strategy useful in the design of

new optimized ligands and prototypes with new molecular architectures composed of two

or more known bioactive pharmacophoric fragments, through the adequate fusion into a

single molecule. The advantage of using molecular hybridization is to activate different

targets by a single molecule, thereby increasing therapeutic efficacy as well as to improve

the bioavailability profile. Molecular hybridization approach is being used in designing

new ligands belonging to different therapeutic categories such as antitubercular,

antithrombotic, antibacterial, antidiabetic, and anticancer.

Applications in medicinal chemistry

Carvalho et al., synthesized a hydrazine series of compounds 1 and 2 through

molecular hybridization of the megazol and the guanylhydrazone derivative. These hybrids

were evaluated for trypanocide activity resulting IC50 = 5.3 µM and IC50 = 63.4 µM.

Kitajima et al., developed a potent nonsulfonylureainsulinotropic agent 2 by hybridizing

KAD-1229 and pioglitazone which demonstrated insulin sensitizing activity. Dardonville

et al., reported a new opioid molecular hybrids by hybridizing fentanyl (an opioid agent)

and agmatine (a ligand of I2-IBS receptor). The hybrid derivative 4 (K i=6.5 nM) presenting

significant increase in the affinity to I2-IBS receptors, compared to fentanyl (Ki=5462 nM).

Capela et al., synthesized primaquine-artemisinin hybrids by hybridizing Artemisinin and

Primaquine. The new hybrid 5 which displayed antimalarial efficacy superior to that of

Abstract

ii

parent compounds. Silva et al., designed and synthesized Phenothiazine N-acylhydrazone

hybrids by using pyrazolyl-4-acylhydrazone and chlorpromazine. This new hybrid

derivative 6 inhibited 100 % arachidonic acid with more than 10 times the parent

compounds, without interfering the ADP induced aggregation.

N

NO2N

S

NN

NH

CH3

N

H

OH

OH S

NN

NH

N

H

OH

OH

1 2

N

O CH3

O

COOH

O NH2

34: n =12

N N

NH

O

n

H2N

NH

O

CH3

O

OO

H3C

H3C

O

HN

NH

N

OCH3

5

NS

NH

N

H

O

H

O

6

Figure 1: New molecular hybrids with different disease targets

Introduction to Hypertension

At present cardiovascular diseases (CVDs) indicates the second cause of worldwide

mortality and morbidity. RAAS mechanism plays an important role in controlling blood

pressure. Renin converts angiotensinogen (dodecapeptide and Liver derived) to angiotensin

I (decapeptide) is the rate-limiting system of this system. Angiotensin I (Ang I) then

converted by angiotensin-converting enzyme to the octapeptide angiotensin II (Ang II).

This small peptide interacts with multiple different systems (two receptors located in the

different target organs, namely AT1 and AT2) and possesses various physiological actions.

Abstract

iii

Angiotensin converting enzyme (ACE), is a pivotal component of the Renin-angiotensin

and aldosterone system (RAAS), playing an important role in maintaining normal blood

pressure and regulation and control of arterial pressure. In recent times, ACE inhibitors are recommended as first line therapy for treatment of

hypertension in children and second line therapy in geriatric patients. The clinical use of

ACE inhibitors is strongly recommended due to their ability in preventing and reversing

functional and structural alterations associated with hypertension. Some of the ACE drugs

in the market are Captopril, Lisinopril, Enalapril, Ramipril, Quinapril, Benazepril and

Alacepril (Figure 2).

Figure 2: ACE drugs in the market

Introduction to Tuberculosis

Tuberculosis (TB) is an ancient chronic infectious disease caused mainly by pathogen

Mycobacterium tuberculosis (Mtb). According to the latest world health organization

(WHO) report there were 8.7 million TB cases, including 1.1 million cases among people

with HIV. In 2011 alone 1.4 million people died because of TB, including half a million

are women and 430,000 people co-infected with HIV. Additionally, the evolution of its

new virulent forms like multi drug resistant tuberculosis (MDR-TB) and extremely drug

resistant tuberculosis (XDR-TB) has become a major threat to human kind. Major

Abstract

iv

obstacles to global control of this infectious disease include the difficulties to detect and

cure sufficient number of cases to interrupt transmission.

In spite of this, there is no specific treatment until 20th century which saw the

development of several antimicrobials leading to the modern day highly effective 6

month’s regimen 2HREZ/4HR3 (two months with four first-line drugs isoniazid,

rifampicin, ethambutol and pyrazinamide followed by a minimum of 4 months of isoniazid

and rifampicin treatment). Second line drugs such as amikacin, kanamycin, capreomycin,

enviomycin, viomycin, ciprofloxacin, levofloxacin, moxiflaxacin, ethionamide,

prothinamide, cyloserin and terizidone known as reserved therapy is used in case of

resistance to first line therapy, extensively drug–resistant tuberculosis (XDR-TB) or

multidrug-resistant tuberculosis (MDR-TB). Recommended treatment regimen is highly

effective and rates of severe adverse reactions are low. However, TB is responsible for

opportunistic secondary infections and co-infections in immune compromised individuals.

N

OHN

NH2

NH

HN

HO

OHN

N

O

NH2

Pyrazinamide (16)Isoniazid (14) Ethambutol (15)

F

O

HO

O

N N

NH

Ciprofloxacin (17)

S

N

NH2

Ethionamide (19)

HN

NH

SQ 109 (18)

O

HO

OHO

N N

N

O

HN

N

OI - A09 (20)

Figure 3: Drugs currently in use and clinical trials for treating tuberculosis

TB drugs currently in use were discovered much before in the seventies and studies

indicate that TB drug research remained still, for almost 40 years. With gained momentum

in antitubercular discovery research, many compounds are presently in advanced stages of

ongoing clinical trials and several of them are in preclinical evaluations. Some of these

drugs were initially discovered as antitubercular agents and have subsequently developed

Abstract

v

as potential TB drugs, which includes the bedaquiline recently approved by the FDA after

40 years. Sutezolid, SQ 109, SQ609, TBA-354, I-A09, Q 203, PA 824 and AZD 5487 are

some drugs in development stages (Figure 3).

Chapter II: Synthesis and evaluation of 2-butyl-4-chloro-1-imidazole

embedded chalcones and pyrazole as Angiotensin

Converting Enzyme (ACE) inhibitors.

Introduction

Naturally occurring flavanoids are having diversified biological activities. In vitro

screening of compounds from ethanobotanical sources have identified Butein (21),

Apigenin (22), and Luteolin (23) (α, β unsaturated ketones) as one of the safe, potent ACE

inhibitory compounds. It is well known that α, β-unsaturated esters and amides (i.e

chalcone) can react with the catalytic nucleophile at the active site of enzyme to give a

Michael type addition product, thus modifying the nucleophile covalently. The unique

pharmacophoric structural feature of this compound is chalcone architecture.

Figure 4: Natural ACE inhibitors with chalcone architecture.

The choice of heterocyclic units arised from clinically used drug molecules as potent

RAAS effectors. Among the several RAAS components, Angiotensin II plays critical

integral role in the pathophysiology of hypertension. Drugs such as Losartan, Eprosartan

(Figure 5) developed as Angiotensin II antagonists, are clinically used in conjunction with

ACE inhibitors for treating hypertension and then cardiovascular diseases.

Abstract

vi

Figure 5: AT1 receptor antagonists

Present work

In present work we made an effort to design new heterocyclic library with drug like

properties, the 2-butyl-4-chloro-1H-imidazole unit of Losartan or Eprosartan and chalcone

architecture of natural Butein were embedded in one molecular frame (Figure 6) to

evaluate their potency as ACE inhibitors.

Figure 6: Design strategy for the development of new ACE inhibitors

Synthesis of 2-butyl-4-chloro-1H-imidazole-5-carbaldehyde building block:

Pentanimidate hydrochloride 27 is generally prepared by passing HCl gas through a

solution of pentanonitrile 26 and MeOH at -15 oC to 0 oC, and stirred at 20 oC for 12 h and

was neutralized with aqueous KOH solution. The toluene solution of the free base was

directly added to glycine suspension in MeOH-H2O without any further treatment. After 15

h, crude reaction mixture was subjected to the modified Vilsmeier reaction by treating it

with POCl3 at -5 oC followed by addition of DMF was heated to 100 oC for 3h. We devised

Abstract

vii

an efficient method to isolate the product from the impurities by exploiting the acid-base

nature of the imidazole formed. The toluene extract having the product in solution was

treated with 10% aqueous KOH and the aqueous portion was carefully acidified with acetic

acid to precipitate a solid that was recrystallized in toluene-cyclohexane solvent mixture

gave 2-butyl-4-chloro-1H-imidazole-5-carbaldehyde 29 a pale yellow solid.(Scheme 1)

Scheme 1: Reagents & conditions (a) i.MeOH. HCl(g), 12 h, 20 oC, ii. Toulene, KOH

(6M): (b) Glycine, MeOH-H2O, 15 h, 40oC: (c) POCl3, DMF, 100 oC, 3 h

The imidazole aldehyde 29 was reacted with methyl iodide (in case of 30a)/ substituted

aryl and heteroaryl benzyl bromides (30b-h) in K2CO3 and DMF at 0°C - RT to give N-

substituted 2-butyl-4-chloro-1-imidazolyl-5-carbaldehyde 30a-h in excellent yields.

(Scheme 2)

Scheme 2

Synthesis of Chalcones

With requisite imidazole derivative 31a in hand, initially, the base catalyzed Claisen-

Schmidt condensation of 2-butyl-4-chloro-1-methyl-1H-imidazoyl-5-carbaldehyde 31a

with various aryl/ heteroaryl acetophenones 32a-aq were reacted in the presence of 10%

aqueous sodium hydroxide in methanol for 3-5 h to gave chalcones 33a-aq in very good

yields (Scheme 3). All the compounds are characterized from 1H-NMR, 13C-NMR and

mass spectral data. The structure of compound 33q confirmed from single X-ray crystal

structure (Figure 7).

Abstract

viii

Scheme 3

Figure 7: ORTEP representation of compound 33q with thermal displacement ellipsoids

drawn at the 30% probability.

Pharmacology: All the fourty three newly synthesized chalcones 33a-aq were screened

for their in vitro ACE inhibitory activity. Among them one derivative 33s found to be

Abstract

ix

most active with % inhibition of 80.61 and it is correlated with natural products and found

to be 100 folds more potent. The most potent compound (E)-3-(2-butyl-4-chloro-1-methyl-

1H-imidazol-5-yl)-1-(1H-pyrrol-2-yl)prop-2-none 33s is also correlated with reference

drugs and shown greater inhibition than standard drugs Enalapril and Lisinopril with %

inhibition of 64.25, 79.54 and less inhibition than Ramipril, Benazepril, Quinapril with %

inhibition of 82.85, 91.58, 91.58.

Compound optimization: Conversion Hit-to-Lead compounds:

Preliminary examination of ACE inhibitory activity revealed that chalcones derivatives

with 2-acetyl pyrrole are found to be more potent than all the derivatives examined.

Scheme 4

In order to further optimize chalcone analogues, we have synthesized a series new

chalcone derivatives 34b-h by reacting acetyl pyrrole 32s with “N” substituted imidazole

derivatives i.e., 31b-h (Scheme 4). Compound 34c is characterized from 1H-NMR by the

appearance as broad singlet at δ 9.73 for NH proton (1H) of pyrrole, trans olefinic protons

appeared at δ 7.57(d, J = 15.8 Hz, 1H), δ 7.47 (d, J = 15.8 Hz, 1H), at δ 5.16 appeared as

singlet for benzylic protons (2H). In ESI-MS spectrum showed peak at m/z 458 [M+H]+ as

molecular ion and confirmed the structure 34c as (E)-3-(2-butyl-4-chloro-1-(3,4,5-

trimethoxybenzyl)-1H-imidazol-5-yl)-1-(1H-pyrrol-2-yl)prop-2-en-1-one.

Pharmacology: All the synthesized compounds were screened for ACE inhibitory activity.

Among them compound 34c is found to be most potent with 84.09% inhibiton, when

Abstract

x

compared with standard drugs Enalapril, Lisinopril, Ramipril, with % inhibition of 64.25,

79.54, 82.85 and less potent than Benazepril, Quinapril with % inhibiton of 91.58, 91.58.

Lead Optimization:

We further continued our efforts to attain most potent ACE inhibitor activity. Synthesis of

chalcones 35(a-b), 35(d-g), 35j, 35(m-n), 35(p-q), 35t, 35(ae-ah), 35(ak-am), 35ao

(Scheme 5) was carried out by reacting aldehyde 31c with various acetophenones 32(a-b),

(d-h), j, (m-n), (p-q),t, (ae-ah), (ak-am), ao. for generation of lead compound. All the

products and their structural characterization were carried out 1H-NMR, 13C-NMR, IR and

mass spectral data. Further It is unambiguously confirmed the structure 35q by single

crystal X-ray diffraction studies (Figure 8).

N

N

Cl

CHO

R' CH3

O

+

N

N

Cl

R'

O

10% aq. NaOH/ MeOH

RT, 3-5 h

32(a-b),(d-h),

j,(m-n), (p-q), t,ae, (af-ah),(ak-am),ao

35(a-b),(d-h),j,(m-n), (p-q), t,

(ae-ah),(ak-am),ao

31c

MeO

MeO

OMeOMe

MeO

MeO

S

NH

O

O

Z

AY

X

35a: X = Y = Z = A = H35b: X = Y = A = H; Z = CH3

35d: X = OCH3; Y = Z= A = H35e: X = Z = A = H; Y = OCH3

35f: X = Y = A = H; Z = OCH3

35g: X = Y = H; Z = A = OCH3

35h: X = H; Y= Z= A = OCH3

XA

35n: X = S; A = H35p: X = S; A = Cl35q: X = S; A = Br

O

F35ae

X

35af: X = O; Y = H35ag: X = O; Y = OCH3

35ah: X = S; Y = H

Y

X

35ak: X= O35al: X= S

35m 35t

35am35ao

R'=

Scheme 5

Abstract

xi

Figure 8: ORTEP representation of compound 35q with thermal displacement ellipsoids

drawn at the 30% probability.

Pharmacology: All the synthesized compounds 35(a-b), (d-h), j, (m-n), (p-q), t, (ae-ah),

(ak-am), ao were screened for ACE inhibitory activity. Structural activity relationship

studies shows ACE inhibition activity is in the order of 35d > 35e > 35f. Among them

compound 35am found to be most active with 100 % inhibiton, compared with standard

drugs Enalapril, Lisinopril, Ramipril, Benazepril, Quinapril with % inhibiton of 64.25,

79.54, 82.85, 91.58, 91.58. Hence (E)-3-(2-butyl-4-chloro-1-(3,4,5-trimethoxybenzyl)-1H-

imidazol-5-yl)-1-(9H-carbazol-2-yl)prop-2-en-1-one 35am is emerged as lead compound.

Synthesis of pyrazoles

Further to obtain compounds containing pharmacophoric pyrazole unit, some

randomly selected chalcones 33(a-b), f, h, k, n, (p-s), v, aa, aj, ak, al, an-ao, aq was

subjected to the condensation reaction with hydrazine hydrate in acetic acid at reflux

temperature for 3-5 h to afford 36(a-b), f, h, k, n, (p-s), v, aa, aj, ak, al, an-ao, aq in

excellent yields (Scheme 6). All the products and their structural characterization were

carried by 1H-NMR, 13C-NMR, IR and mass spectral data. The structure of compound 36p

is confirmed by single crystal X-ray diffraction studies (Figure 9).

Abstract

xii

Scheme 6

Figure 9: ORTEP representation of compound 36p with thermal displacement ellipsoids

drawn at the 30% probability.

Pharmacology: All the synthesized pyrazole analogs were screened for ACE inhibitory

activity and are found to be less active than chalcone analogs.

Conclusion In conclusion, we have described an efficient synthesis and evaluation of ACE

inhibitory activity of novel 2-butyl-4-chloro-N-substituted imidazole derived chalcones

and pyrazoles. The chalcones were prepared in excellent yields through Claisen–Schmidt

Abstract

xiii

condensation of 2-butyl-4-chloro-N-substituted imidazole-5-carbaldehyde 31a-h with

aryl/heteroaryl methyl ketones 32a-aq in the presence of 10% aqueous NaOH in methanol.

These chalcones were further derivatized with hydrazine hydrate in acetic acid to give

pyrazole analogues. In preliminary evaluation of imidazole derived chalcones 33a-aq,

resulted one derivative 33s was found to be most potent. Compound 33s is taken for lead

generation by varying substitutions at nitrogen of imidazole ring 34b-h, among them N-

substituted 3,4,5-trimethoxybenzyl substituted chalcone 34c (84.09 %) resulted with

greater inhibition compared to that of standard drugs. Compound 34c is taken for lead

optimization and synthesized chalcones 35(a-b), (d-h), j, (m-n), (p-q), t, (ae-ah), (ak-am),

ao, by varying different aryl and heteroaryl methylketones, the chalcone 35am emerged as

lead compound with 100 % inhibition. Among all the chalcones derivatives 35am (IC50 0.5

nM) found to be most potent than all the standard drugs Enalapril (IC50 745 nM), Lisinopril

(IC50 298 nM), Ramipril (IC50 117 nM), Benazepril (IC50 326 nM), Quinapril (IC50 433

nM).

Chapter III: Synthesis and evaluation of novel 2-butyl-4-chloro-imidazole

derived peptidomimics as Angiotensin Converting Enzyme

(ACE) inhibitors.

Introduction

During the course of evolution, nature has developed a vast number of peptides in all

living species that display diversity of structure and biological effects, such as hormonal

and enzyme-controlling activity, communication between cells, and participation in host

defence. Natural occurring Dolastatin (37) a linear peptide containing three unique amino

acid units was originally isolated from Indian Ocean sea hare Dolabella auricularia with

antineoplastic activity. Geodiamolides(A-F) (38) are a family of bioactive marine sponge-

derived cyclodepsipeptides exhibits in vitro cytotoxicity against human cancer cell lines.

Many efforts has been devoted recently to synthesize of peptidomimetics were designed to

overcome the therapeutic deficiencies of peptides. Despite of these generic advantages of

peptides more than 100 peptide-based drugs have already reached the market.

Abstract

xiv

Present work

In present work, novel dipeptidomimics and tripeptidomimics derived 2-butyl 4-chloro

imidazole have been designed and synthesized using molecular hybridization approach by

integrating two pharmacophoric fragments i.e from AT1 receptor antagonist and ACE

inhibitor, (Figure 10) could generate potential new scaffolds for ACE inhibitory activity.

Figure 10: Design strategy for the development of ACE inhibitors

Synthesis of Imidazolyl based dipeptidomimics

The preparation starting material 29 and 31a has been described in Chapter II .

Oxidation of the 31a with KMnO4 in acetone and water (1:1) obtain acid 39 in 75 % yield.

Coupling of L-aminoacid methyl esters of 40a-k with 39 using coupling reagent1-Ethyl-3-

Abstract

xv

(3-dimethylaminopropyl) carbodiimide (EDC), Hydroxybenzotriazole (HOBT), N,N-

diisopropylamine as base in dichloromethane to obtain 41a-k in 70-79% yield.

Deprotection methyl ester of 41a-k with lithium hydroxide in THF, methanol and water

(3:1:1) stirred for 30 min to obtain 42a-k in 95-98 %yield.(Scheme 7) All the compounds

were characterized by 1H-NMR, 13C-NMR, IR and Mass spectral data. Compound 42b

(Figure 11) is unambiguously confirmed the structure and stereochemistry by single

crystal X-ray diffraction studies.

Scheme 7

Figure 11: ORTEP representation of compound 42b with thermal displacement ellipsoids

drawn at the 30% probability.

Synthesis of Imidazole-Proline based tripeptidomimics

Abstract

xvi

For the synthesis of tripeptide, initially proline 43 was protected with Boc

anhydride in the presence of 4N sodium hydroxide for 6h stirring followed by

neutralization with potassium bisulphate extracted with ethyl acetate to give 44 in 95%

yield. The compound 44 was coupled with methyl ester protected L-aminoacids 45a-j in

presence of EDC, HOBT, N,N-diisopropylamine in dichloromethane at 0 °C -RT for 6h to

afford 46a-j in excellent yields. Boc was deprotected with TFA in DCM at 0 °C for 30 min

to obtain 47a-j in good yields (Scheme 8) and which was used as such in the next step

without any characterization and further purification.

Scheme 8: Reagents and conditions: a) Boc anhydride, 4N NaOH; b) EDC, HOBT,

DIPEA, DCM, 0°C-RT, 6h; c) TFA.DCM. 0°C-RT, 30 min.

Synthesis of 2-butyl-4-chloro-1-(3-methylbenzyl)-1H-imidazole-5-carboxylic acid:

The Synthesis of starting material 31b has been reported in (Chapter II) . Oxidation of the

31b with KMnO4 in acetone and water (1:1, % v/v) refluxed for 18h, and then neutralized

with conc. HCl gave acid 48 as acid in 73% yield. Compound 48 was coupled with 47a-j

in presence of EDC, HOBT, and N,N-diisopropylamine in dichloromethane at 0 °C–RT for

6h to afforded 49a-j in 65-73 % yields. Deprotection methyl ester of 49a-j with lithium

hydroxide in THF, Methanol and water (3:1:1) stirring for 30min to obtain 50a-j in good

yields (Scheme 9). Compound 50h was characterized by 1H-NMR spectrum by the

appearance of multiplet at δ 7.01-7.33 (10H) for two phenyl rings, at δ 5.27 (J = 15.8 Hz,

1H) and δ 5.15(J = 15.8 Hz, 1H) as two doublets for benzylic protons (2H) attached to

imidazole ring, at δ 4.66-4.69(1H) and δ 4.36-4.45(1H) appeared as two multiplets for (-

CH2) protons of phenylalanine. In the ESI-MS spectrum a peak at m/z 537[M+H]+

Abstract

xvii

confirmed the structure 50h as (S)-2-((S)-1-(1-Benzyl-2-butyl-4-chloro-1H-imidazole-5-

carbonyl) pyrro -lidine-2-carboxamido)-3-phenyl propanoic acid.

Scheme 9: Reagents and conditions: a) KMnO4, acetone: H2O 1:1, 18h, reflux; b) (i)47a-

j , EDC, HOBT, DIPEA, DCM, 0 °C-RT, 6h; (ii) LiOH, THF:MeOH:H2O, (3:1:1), 30 min.

in two steps.

Pharmacology: The synthetic imidazole derived dipeptide and tripeptide analogues 40i,

40k, 41e, 41h, 41i, 49e, 49g, 50a, 50e and 50h displayed good ACE inhibitory activity

greater than 62% inhibition and were evaluated for dose response study at different

concentrations ranging from 1µM-100nM and showed with IC50 0.647, 0.531, 1.12, 0.657

0.1, 0.350, 0.304, 0.398, 0.477 and 0.347µM respectively. Among them compound 17i (0.1

µM) found to be most potent compared to standard drug lisinopril.

Conclusion

In conclusion we have synthesized novel imidazole derived dipeptide and tripeptide

peptidomimics by the reaction of imidazolecarboxylic acid with the appropriate L-amino

acids. All the new derivatives were evaluated for their invitro ACE inhibitory activity

resulted ten analogues 40i, 40k, 41e, 41h, 41i, 49e, 49g, 50a, 50e and 50h displayed

good ACE inhibitory activity. Among them (S)-2-(2-butyl-4-chloro-1-methyl-1H-

imidazole-5-carboxamido)-3-phenylpropanoic acid 41i (IC50 0.1µM) emerged as most

active carboxylic acid ACE inhibitor with minimal toxicity comparable to standard drug

lisinopril.

Abstract

xviii

Chapter IV: Synthesis and evaluation of novel 2-hydroxypyrrolo

benzodiazepine-5,11-dione analogues as potent

Angiotensin Converting Enzyme (ACE) inhibitors.

Introduction

Naturally occurring alkaloids, pyrrolo [2,1-c][1,4] benzodiazepines (PBDs) isolated

from various Streptomycin species (a class of biologically active compounds with good

pharmacological profile). These natural and synthetic PBDs (Figure 12) exert anticancer

activity through their interaction towards protein sequences to interfere and are currently in

clinical development.

Figure 12: Natural PBDs

Present work

Clinical ACE drugs found to possess proline or modified proline type architecture as a

critical pharmacophore responsible for exhibiting ACE inhibitor activity. With growing

interest in use of pyrrolo [2,1-c] [1,4] benzodiazepine (PBD) ring system as a potential

pharmacophoric fragment. A series of novel 10-substituted 2-hydroxy pyrrolo

benzodiazepine-5,11-diones designed through structure based rational hybridization

approach, Benazepril (ACE inhibitor)with fused seven member cyclic ring has structural

similarity to basic benzodiazepine architecture. This rekindled our curiousness to inculcate

a design strategy (Figure 13) integrating Proline fragment in benzodiazepine architecture

with appended lipophilic functionality.

Abstract

xix

Figure 13: Structure based rational design strategy

Synthetic approach for the preparation of hydroxypyrrolo [2,1-c][1,4] benzodiazepine-

5,11-dione 58a-w was presented in Scheme 10. Isotonic anhydride 54 on

Cyclodehydration with readily available (2S,4R)-4-hydroxypyrrolidine-2-carboxylic acid

55 in DMSO resulted (2R,11aS)-2-hydroxy-2,3-dihydro-1H-benzo[e]pyrrolo [1,2-

a][1,4]diazepine-5,11(10H,11aH)dione 56 in excellent yield.

Scheme 10

Abstract

xx

This hydroxypyrrolobenzodiazepine diones frame unit 56, was then alkylated with

series of alkyl or benzyl bromides 57b-w (methyl iodide in case of 57a), in the presence of

K2CO3 in DMF to gave 10-substituted (2R,11aS)-2-hydroxy-2,3-dihydro-1H-benzo [e]

pyrrolo [1,2-a][1,4] diazepine-5,11(10H,11aH) diones 58a-w in excellent yields.

Compound 58g unambiguously confirmed the structure and stereochemistry by single

crystal X-ray structure (Figure 14).

Figure 14: ORTEP representation of compounds 58g with thermal displacement ellipsoids

drawn at the 30% probability

Conclusion

All newly synthesized compounds 58a-w were screened ACE inhibitory activity.

Among them one compound (2R,11aS)-10-((4-bromothiophen-2-yl)methyl)-2-hydroxy-

2,3-di hydro-1H-benzo[e]pyrrolo[1,2-a][1,4] diazepine-5,11(10H,11aH) dione, 58v (IC50:

0.272 µM) emerged as most active non-carboxylic acid ACE inhibitor with minimal

toxicity comparable to clinical drugs Lisinopril, Benazepril and Ramipril.

Chapter V &

Section A

: Synthesis of dihydrobenzofuran tethered pyridines

and dihydro-6H-quinolin-5-ones from β-enaminones

as potent inhibitors of Mycobacterium tuberculosis

Introduction

The prevalence and diversity of polysubstituted aromatic nitrogen heterocyclic viz:

pyridine, quinolinones, found in natural products and used in medicinal chemistry

continues to fuel the development of newer methods and strategies for their synthesis.

Abstract

xxi

Notable among natural products and other successful synthetic drug candidates having

2,3,6-substituted pyridine central core unit (Figure 15).

Figure 15: Natural and synthetic drug candidates having pyridine core unit

In an effort to develop new heterocyclic library with drug like properties, we thought to

synthesize a series of analogues containing both pyridine and dihydrobenzofuran moieties

in one molecular frame. A promising approach for the preparation of this structurally

related pyridine-dihydrobenzofuran conjugates would be through multicomponent

reactions (MCR) and are screened for antitubercular activity.

Present work

In recent years, among the lanthanide catalysts, cerium (III) chloride has emerged

as a very cheap, efficient, green reagent and is able to catalyze various selective chemical

transformations and cyclization. In most cases, the activity of CeCl3 can be increased in

combination with NaI. The successful utility of cerium (III) in reactions originated from

1,3-dicarbonyls, prompted us to investigate its applicability in one-pot condensation of β-

enaminones with 1,3-dicarbonyls and ammonium acetate.

Synthesis of Acetyl dihydrobenzofuran derivatives

Synthesis of acetyl dihydrobenzofuran 68a-b was achieved according to the reported

procedure. Allylation of 2-hydroxy-4-methylacetophenone 62a and 5-fluoro-2-hydroxy

acetophenone 62b with allyl bromide, K2CO3 as base gave 63a-b in good yields. Claisen

rearrangement of 63a-b under thermal condition at 180 °C to obtain 64a-b as major

product, then acid catalyzed regioselective cyclisation of 64a-b in 1,4 dioxane heated at

100 °C for 4h gave acetyl dihydrobenzofuran derivatives 65a-b in 70-73 % yields (Scheme

11).

Abstract

xxii

Scheme 11

1-(2,4-dimethyl-2,3-dihydrobenzofuran-7-yl)ethanone 65a and 1-(5-fluoro-2-methyl-2,3-

dihydro benzofuran-7-yl)ethanone 65b were converted to β-enaminones 66a-b by

refluxing with dimethylformamide dimethylacetal (DMF-DMA) in xylene (Scheme 12) .

Scheme 12

The structure of 66b is characterized from NMR, mass spectra and confirmed by single

crystal X-ray diffraction analysis (Figure 16).

Figure 16: ORTEP representation of compounds 66b with thermal displacement ellipsoids

drawn at the 30% probability.

Initially β-enaminone 66a was selected to react with 1,3-acyclic dicarbonyl 67a-f and

ammonium acetate catalyzed by CeCl3.7H2O-NaI to gave 68aa-af (Scheme 13). Similarly

66b reacts with 67a, 67d to obtain 68ba and 68bd in good yields. The compounds 68aa

Abstract

xxiii

and 68ae are unambiguously confirmed the structure by single crystal X-ray diffraction

studies (Figure 17).

Scheme 13

Figure 17: ORTEP representation of compounds 68aa and 68ae with thermal

displacement ellipsoids drawn at the 30% probability.

Reaction of β-enaminone 66a with 1,3-cyclic diketones 67g-k and ammonium acetate

in the presence of CeCl3.7H2O-NaI under optimized conditions afforded 68ag-ak in 85 %

yield. Similarly β-enaminones 66b reacts with 67g-i to give 68bg-bi respectively in good

yields. Other hand, to achieve further diversity, β-enaminones 66a & 66b were condensed

with 1,3-indanedione 67k under standard one-pot protocol to result novel analogs of

indeno[1,2-b]pyridin-5-ones 68ak & 68bk in very good yields.(Scheme 14)

Abstract

xxiv

Scheme 14

Furthermore the current method is easy to perform and broad in scope to generate

novel library of substituted dihydroquinolin-5-one motifs bearing dihydrobenzofuran.

2-Chloromethyl-6-(2,4-dimethyl-2,3-dihydro benzofuran-7-yl) nicotinic acid ethyl ester

68ae was treated with Pyridine, DMAP, N-phenyl piperazine 69 in acetonitrile at room

temperature (Scheme 15). Ethyl 6-(2,4-dimethyl-2,3-dihydrobenzofuran-7-yl)-2-((4-

phenylpiperazin-1-yl)methyl nicotinate 70 obtained in 77 % yield.

Scheme 15

Conclusion

In conclusion, we have developed an efficient cerium (III) catalyzed protocol for one-pot

regioselective synthesis of novel dihydrobenzofuran tethered pyridines and dihydro-6H-quinolin-

5-ones employing β–enaminones 66a-b and acyclic & cyclic 1,3-dicarbonyls as novel variants of

Bohltzmann-Rahtz substrate, and ammonium acetate in refluxing 2-propanol. Mechanistically,

the reaction proceeds through sequential Michael addition-cyclodehydration and elimination

Abstract

xxv

reactions. All the synthesized compounds were screened against Mycobacterium tuberculosis.

Among them the compounds 68ah, 68bh, 68ai and 68i are found to be most potent anti

tubercular agent with MIC 3.12 µg/mL.

Chapter V &

Section B

: Synthesis of 6-substituted 2-picolines from aryl /

heteroaryl-β-enaminones and meldrum’s acid

Introduction

Systematic explorations towards drug discovery 6-substituted-2-picolines 71-73

(Figure 18) play an important part in the development of potent bioactive derivatives. SIB-

1893 (71) & MPEP (72) are the first drugs developed to act as a selective antagonist for the

metabotropic glutamate receptor subtype mGluR5 with anticonvulsant and neuroprotective

effects following acute brain injury and reduced glutamate release. The substituted

picolines are mostly prepared through metal catalyzed aromatic coupling reactions. These

methods use palladium and other metal catalysts and each have limitations in applicability

for library generation and medicinal applications.

Figure 18: Synthetic drugs for metabotropic glutamate receptor subtype mGluR5

Present work

The substituted picolines are mostly prepared through metal catalyzed aromatic

coupling reactions. These methods use palladium and other metal catalysts and each have

limitations in applicability for library generation and medicinal applications. We herein

report one pot efficient CeCl3.7H2O-NaI catalyzed regioselective conversion of

β-enaminones to novel 6-substituted 2-picolines through multicomponent reaction (MCR)

and evaluate for antitubercular activity.

Abstract

xxvi

The condensation enaminones 74a-l meldrum’s acid 75 CeCl3.7H2O-NaI and NH4OAc

in i-PrOH was refluxed for 4h the mixture was cooled to RT, the solid precipitate was

filtered and washed (cold i-PrOH) gave 76a-l as syrups (Scheme 16). Compound 76f was

characterized from the 1H-NMR spectrum by the appearance of singlet at δ 2.59 (3H) for

methyl group attached to pyridine, δ 7.92 (J = 8.3 Hz, 1H) appeared as triplet for 4-H of

pyridine, at δ 7.56 (J = 7.5 Hz, 1H) as doublet for 3-H of pyridine, at δ 7.01 (J = 7.5 Hz,

1H) as doublet for 5-H of pyridine, tertiary carbon proton of dihydrobenzofuran ring

appeared as multiplet at δ 4.97-5.05 for (1H) and secondary carbon protons of

dihydrobenzofuran appeared as doublet of doublet at δ 3.33 and δ 2.83 for (2H) In ESI-MS

spectrum a peak at m/z 244 [M+H]+ as molecular ion, confirmed the structure 76f 2-(5-

Fluoro-2-methyl-2,3-dihydrobenzofuran-7-yl)-6-methyl pyridine.

Scheme 16

Conclusion

In summary, we have developed a cerium (III) catalyzed protocol for one-pot

regioselective synthesis of novel 6-substituted 2-picolines 76a-l from β–enaminones

74a-l, meldrum’s acid 75 and ammonium acetate. The reaction proceeds through

sequential Michael addition-cyclodehydration and elimination reactions. It is likely that

the novelty of this method combined with its operational simplicity will make it

Abstract

xxvii

attractive for the construction of environmentally benign, low cost and biologically

relevant compounds. All the compounds are evaluated antitubercular activity and found

to be less potent than standard drugs.

Chapter VI

& Section A

: Synthesis and antimicrobial evaluation of novel 1-

benzyl 2-butyl-4-chloro imidazole embodied 4-

Azafluorenones

Introduction

4-Azafluorenones (5H-indeno[1,2-b]pyridin-5-one) are the naturally occurring

alkaloids isolated from the root of the plant Polyalthia debilis belonging to the family of

Annonaceae, the root water decoction of which has been traditionally used for treatment of

antimicrobial infections (Figure 19).

Figure 19: Examples of bioactive natural 77-80 and synthetic 81 Azaflourenone.

Present work

The design strategy adopted here for library generation was based on most recent

molecular hybridization approach. Considering antimicrobial Onychine 77 as a basic

bioactive unit of natural 4-azafluorenone, the modifications were introduced on nucleus

through hybridization with chosen imidazole pharmacophore (1-benzyl-2-butyl-4-

chloroimidazole) and evaluated for their antimicrobial activity (Figure 20).

Abstract

xxviii

Figure 20: Illustration of design strategy for library generation.

A series, imidazole embodied 4-azafluorenone analogues 84a-o were prepared through

condensation between 1-benzyl-2-butyl-4-chloroimidazole carbaldehyde 31b (synthesis of

31b is described in Chapter II ) aryl/ heteroaryl methyl ketones 83a-o, 1,3-indanedione 82

and ammonium acetate in DMF refluxed for 3- 4 h gave 84a-o in 77-86 % yields as

yellow solids.(Scheme 15) Structure of compound 84h is unambiguously confirmed by the

single crystal X-ray diffraction studies (Figure 21).

N

N

Cl

H

O

O

O

R' CH3

ONH4OAc

DMF, reflux,3-4h.

N

N

Cl

N

O

R'82 83a-o 84a-o

Y

X Z

84a: X=Y=Z=H

84b: X=Z=H;Y=CH3

84c: X=Z=H;Y=Br

84d: X=Z=H;Y=OCH3

84e: X=Z=H;Y=NO2

84f: X=Y=Z=OCH3

SSBr O

NBr A84m: A=O84n: A=S

N S

O

O

84g 84h 84i 84j 84k

84l

R'=

31b

84o

Scheme 15

Abstract

xxix

Figure 21: ORTEP representation of compound 84h with thermal displacement ellipsoids

drawn at the 30% probability.

Conclusion

In conclusion we have synthesized a novel Azaflourenones 84a-o through one pot three

component condensation of imidazole aldehyde, 1,3-indanedione and ammonium acetate

under reflux condition. Screening of all new analogs for antimicrobial activity resulted one

compound 84k has pronounced activity with higher zone of inhibition against

Staphylococcus aureus, Pseudomonas aeruginosa, Klebsiella pneumoniae, Aspergillus

flavus and Candida albicans than standard drugs. This results demonstrated potential

importance of molecular hybridization in the development of 84k as potential

antimicrobial agent.

Chapter VI

& Section B

: Synthesis of novel 2-(trifluoromethyl)phenothiazine

[1,2,3]triazoles and [1,2,3]triazole 1-Adamantyl

acetamide hybrids as potent inhibitors of

Mycobacterium tuberculosis

Introduction

Phenothiazine: In early days of organic chemistry, the phenothiazine and its derivatives

played a prominent role in pharmacology and biomedicine. Classical phenothiazine deals

with promising anticancer, antibacterial, antiplasmids, multidrug resistance (MDR) and

potential treatment in Alzheimer’s and Creutzfeldt–Jakob diseases.

Abstract

xxx

Figure 22: Phenathiazine based drug candidates

Phenothiazine based drug candidates (Figure 22) for treating neurodegenerative

disorders were also effective inhibiting M. tuberculosis. Chlorpromazine, trifluoperazine

(TPZ) and thioridazine are few found to act in synergy with regular antibiotics to which M.

tuberculosis is susceptible.

Adamantane: An emerging field with respect to the application of adamantane derivatives

gained significance in medicinal chemistry. Adamantane is “lipophilic bullet” The

adamantane modifications were chosen to enhance Lipophilicity and stability of the drugs,

thereby improving their pharmacokinetics. Several adamantane derived drugs are available

in the market (Figure 23) and also they show diverse biological properties such as

antiviral, antibacterial, antitubercular, antifungal, anti-inflammatory, antidiabetic, and other

medicinal properties.

Figure 23: Drugs containing adamantane nucleus

Abstract

xxxi

The triazole scaffolds are found in a number of biologically active compounds

exhibiting anti-HIV, antibiotics, antiviral, and antibacterial activities. Besides this, triazole

based antitubercular agents (Figure 24) may be regarded as a new class providing truly

effective lead candidates which are reported to inhibit bacteria. Among them I-A09 is

presently in clinical trials.

Figure 24: Triazole based antitubercular agents

Present work

It is therefore of our interest to integrate both 2-(trifluoromethyl)-10H-

phenothiazine, 1-adamantylamine and triazole pharmacophoric fragments in one molecular

platform to generate a new scaffold for antitubercular evaluations.(Figure 25)

Figure 25: Design strategy for new [1,2,3]-triazole hybrids

Abstract

xxxii

Synthesis of 2-(trifluoromethyl) phenothiazine-[1,2,3]triazoles hybrids

Synthesis of 2-(trifluoromethyl)phenothiazine-[1,2,3]triazole hybrids has been carried

out by the commercially available starting material 2-(trifluoromethyl)-10H-phenothiazine

(96) and chloroacetyl chloride were refluxed in toluene to afford the compound (97) in

quantitative yield. This product (97) upon treatment with sodium azide in the presence of

tetra-n-butylammonium bromide produced 2-azido-1-(2-(trifluoromethyl)-10H-pheno

thiazin-10-yl)ethanone (98) in good yield, followed by Huisgen’s (3+2) cycloaddition

reaction with different terminal alkynes (99a-v ) in the presence of CuSO4 catalyst, sodium

ascorbate in t-butanol and water (1:1, % v/v) at RT gave 1,2,3 triazole hybrids (100a-v) in

excellent yields(Scheme 16).

Scheme 16: Reaction conditions: (i) Chloroacetyl chloride, toulene, reflux, 6h, (ii) NaN3,

tetra-n-butylammonium bromide, dichloromethane: H2O (1:1), (iii) CuSO4.5H2O, sodium

ascorbate, t-BuOH, H2O (1:1), 1-2 h, RT

Compound 100m was characterized from 1H-NMR by the appearance of acetylic CH2

protons at δ 5.55 as broad doublet, at δ 3.81 appeared as singlet for 4-OCH3 protons, at δ

Abstract

xxxiii

2.42 appeared as singlet for 2-CH3 protons. In ESI-MS spectrum a peak at m/z 497 [M+H]+

indicates molecular ion, confirmed the structure 100m as 2-(4-(4-methoxy-2-

methylphenyl)-1H-1,2,3-triazol-1-yl)-1-(2-(trifluoromethyl)-10H-phenothia-zin-10-yl)

ethanone.

Synthesis of [1,2,3]-triazole 1-Adamantyl acetamide hybrids

Synthesis of novel Adamantyl-[1,2,3]triazole acetamide hybrids were synthesized

from 1-Adamantanyl amine 101 was made react with chloroacetyl chloride and potassium

carbonate in dichloromethane to gave N-(Adamantyl)-2-chloroacetamide 102 in very good

yield. Reaction of 102 with sodium azide in the presence of tetra-n-butylammonium

bromide in dichloromethane and water (1:1, % v/v) to afford N-(Adamantyl)-2-

azidoacetamide (103) in excellent yield. The Azide 103 was reacted with various terminal

alkynes 99 a-h, k-q, s-x in the presence of CuSO4 catalyst, sodium ascorbate in t-butanol

and water (1:1, % v/v) to produce 1,2,3-triazole hybrids 104 a-h, k-q, s-x in excellent

yields (Scheme 17).

Scheme 17:Reaction conditions: (i) Chloroacetyl chloride, K2CO3,DCM, reflux, 12h, (ii)

NaN3, tetra-n-butyl ammonium bromide, DCM:H2O (1:1) 12h. (iii) CuSO4.5H2O, sodium

ascorbate, t-BuOH:H2O (1:1), 1-2 h, RT.

Compound 104l was characterized from 1H-NMR spectrum by the appearance of broad

singlet at δ 5.83 for NH proton (1H), at δ 4.99 appeared as singlet for acetylic –CH2

protons (2H), at δ 3.87 appeared as singlet for aromatic –OCH3 protons (3H), at δ 2.06

appeared as broad singlet for tertiary carbon protons (3H) of adamantyl ring, at δ 1.96 a

broad singlet for secondary carbon protons (6H) attached to quaternary carbon, at δ 1.65

appeared as broad singlet for secondary carbon protons (6H) attached to tertiary carbon. In

ESI-MS spectrum a peak at m/z 367 [M+H]+ confirmed the structure 104l as N-(1-

Adamantyl)-2-(4-(3-methoxyphenyl)-1H-1,2,3-triazol-1-yl)acetamide.

Abstract

xxxiv

Pharmacology: All newly synthesized compounds 2-(trifluoromethyl) phenothiazine-

[1,2,3]triazoles hybrids 100a-v and [1,2,3]triazole 1-Adamantyl acetamide hybrids 104 a-

h, k-q, s-x were screened for in vitro activity against MTB, shown with MIC ranging from

3.12-50.0 µg/mL. Among all the compounds evaluated seven compounds 100c, 100l, 100o,

104d, 104g, 104q with MIC 6.25 µg/mL. Compound 104u adamantane triazole appended

with phenanthrene, resulted the most active antitubercular agents with MIC 3.12 µg/mL

against M. tuberculosis H37Rv (MTB).

Conclusion

In conclusion we have synthesized a series of novel 2-(trifluoromethyl)

phenothiazine-[1,2,3]-triazoles 100a-v and [1,2,3]-triazole1-Adamantyl acetamide hybrids

104 a-h, k-q, s-x by using Huisgen’s (3+2) cycloaddition reaction between azide and

alkynes in presence of copper sulphate and sodium ascorbate (Click reaction). Evaluation

of all the new hybrids against M. tuberculosis H37Rv (Mtb) and cytotoxicity revealed that

seven compounds 100c, 100l, 100o, 104d, 104g, 104q with MIC 6.25 µg/mL and 104u

with MIC 3.13 µg/mL are the most potent antitubercular agents and with good selectivity

index >15.