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CHAPTER – 4

IN VITRO STUDY OF NOVEL SULFA

HYDRAZONE SUBSTITUTED 4-(3H)-

QUINAZOLINONE HETEROCYCLIC

DERIVATIVES

Chapter -4

Sardar Pate l Univers i ty 229

In this chapter the antibacterial activity of twenty

screened substituted quinazolinone derivetives against

bacterial strains Escherichia coli, Pseudomonas aeruginosa,

Bacillus megaterium and Proteus vulgaris are discussed and

were performed using Agar cup method.

4.1 INTRODUCTION :

The ever-growing resistance to antibiotics leads to

continuous screening for the new biologically effective

substances of natural or synthetic origin [1]. Quinazolinone

derivatives are a subject of increasing interest in bioorganic

and coordination chemistry. A sustained research activity

has been devoted to quinazolinone derivatives, due to their

successful multiple applications as diagnostic tools in

biomedical tribulations like infection and diseases.

Infectious diseases caused by bacteria affect millions of

people worldwide. Rigorous and systematic programs to

discover and develop new antibiotics have been driven to a

considerable extent by the development of resistance by

these organisms to the drugs commonly used against them.

The quinazoline skeleton is one of the ever used as

biologically active molecules since last several decades. In

particular, 4(3H)-quinazolinone derivatives display a broad

range of biological properties such as antihypertensive [2,

3], CNS depressant [4, 5], antitumor, analgesic and anti-

inflammatory [6]. Moreover, 4(3H)-quinazolinone derivatives

indicates the antibacterial and antifungal activities [7]. This

class of compounds also shows the growth inhibition

against Staphylococcus aureus bacteria [8]. Our main focus

is to evaluate the quinazolines derivative for bacteriostatic

effect on multi-drug resistance strains of Staphylococcus

aureus.

Chapter -4


Looking to the vital role of this class of compounds in

many infections and diseases, it was planned to explore the

field of novel anti-infective agents. Thus, the present work

comprises in vitro study of twenty selected substituted

quinazolinone derivetives against Bacillus megaterium,

Escherichia coli, Pseudomonas aerugenosa and Proteus

vulgaris. Besides that the compounds were also analyzed for

their response against the Methicillin-Resistant

Staphylococcus Aureus (MRSA) strains. MRSA is a bacterium

responsible for infections in humans. It may also be

referred to as multiple-resistant Staphylococcus aureus.

4.2. BRIEF HISTORY OF ANTIMICROBIAL SCREENING :

Infection is a major cause of human disease and

skilled management of antimicrobial drugs is of the prime

importance. The term chemotherapy is used for the drug

treatment of parasitic infections in which the parasites

(viruses, bacteria, protozoa, fungi, and worms) are

destroyed or removed without injuring the host. Many

substances that we now know to possess therapeutic

efficacy were first used in the distant past. The Ancient

Greeks used male fern, and the Aztecs chenopodium, as

intestinal anthelmintics. The Ancient Hindus treated leprosy

with chaulmoogra. For hundreds of years moulds have been

applied to wounds. Despite the introduction of mercury as a

treatment for syphilis (16 th century), and the use of

cinchona bark against malaria (17 th century), the history of

modern rational chemotherapy did not begin until Paul

Ehrlich developed the idea from his observation that aniline

dyes selectively stained bacteria in tissue microscopic

preparations and could selectively kill them. He invented

the word ‘chemotherapy’ and in 1906 he wrote : “In order to

use chemotherapy successfully, we must search for

Chapter -4


substances which have an affinity for the cells of the

parasites and a power of killing them greater than the

damage such substances cause to the organism itself… This

means… we must, learn to aim with chemical substances.”

The antimalerials pamaquin and mepacrine were

developed from dyes and in 1935 the first sulphonamides,

linked with a dye (Prontosil), was introduced as a result of

systematic studies by Domagk. The results obtained with

sulphonamides in puerperal sepsis, pneumonia and

meningitis were dramatic and caused a revolution in

scientific and medical thinking. In 1928, Fleming

accidentally discovered the long-known ability of Penicill ium

fungi to suppress the growth of bacterial cultures. In 1939,

principally as an academic exercise, Florey and Chain

undertook an investigation on antibiotics, i.e. substances

produced by microorganisms that are antagonistic to the

growth or life of other microorganisms. They prepared

penicillin and confirmed its remarkable non-toxicity. When

the preparation was administered to a policeman with

combined staphylococcal and streptococcal septicemia,

there was dramatic improvement; unfortunately the

manufacture of penicillin (in the local Pathology Laboratory)

could not keep pace with the requirements (it was also

extracted from the patient’s urine and re-injected); it ran

out and the patient later succumbed to infection.

Subsequent development thoroughly demonstrated the

remarkable therapeutic efficacy of penicillin.

4.3 CLASSIFICATION OF ANTIMICROBIAL DRUGS :

Antimicrobial agents may be classified according to the

type of organism against which they are active.

• Antibacterial drugs.

• Antiviral drugs.

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• Antifungal drugs.

• Antiprotozoal drugs.

A few antimicrobials have useful activity across several

of these groups and they are known as broad spectrum

antibiotics. For example, metronidazole inhibits obligate

anaerobic bacteria (such as Clostridium perfringens) as well

as some protozoa that rely on anarobolic pathways (such as

Trichomonas vaginalis).

Antimicrobial drugs have also been classified broadly

into :

• Bacteriostatic : Those that act primarily by arresting

bacterial multiplication (groth), such as

sulphonamides, tetracyclines and chloramphenicol.

• Bactericidal : Those which act primarily by killing

bacteria, such as penicillins, cephalosporins,

aminoglycosides, isoniazide and rifampicin.

4.4 EVALUATION TECHNIQUES :

The following conditions must be met for the screening

of antimicrobial activity :

• Aseptic/sterile environment should be maintained.

• Mandatory conditions should be provided for the

growth of microorganisms.

• There should be an intimate contact between test

organisms and the substance to be evaluated for its

activity.

• Conditions should be maintained same throughout the

study.

Various methods have been used from time to time by

several workers to evaluate the antimicrobial activity [9-11].

• Turbidometric method.

• Agar streak dilution method.

Chapter -4


• Serial dilution method.

• Agar diffusion method.

Agar diffusion method is again sub classified into three

types:

• Agar cup method.

• Agar ditch method.

• Paper disc Method.

However, in the present work Agar cup method was used.

4.4.1 Agar cup method :

When an antibiotic is added in agar cup (make in

medium previously inoculated with the test organism), the

radial diffusion of an antibiotic through the agar produces a

concentration gradient. Test organism is inhibited at the

minimum inhibitory concentration (MIC), giving rise to a

clear zone on inhibition. Greater the MIC, larger is the zone

of inhibition.

4.4.2 Advantages of agar cup method :

Top agar also known as soft agar or seed agar having

concentration of 0.5% is inoculated with the test organism

and then overlaid on the surface of sterilized nutrient agar

plate. The agar overlay method gives better results due to

the following reasons

• Since it has less concentration of agar (0.5%) it

permits faster diffusion of antibiotics.

• Organism can obtain nourishment from both the top

agar as well as the base agar.

• Since the base agar is free of any inoculum, the

zone of inhibition is sharply defined and clearly

measured.

• The zone of inhibition can be measured clearly as

compared to Paper disc and Agar ditch method.

• The growth of the microorganism in the agar cup

method is uniform in the cups of control, standard

Chapter -4


drug and the antibiotic to be tested, while in the

case of ditch method the growth depends on the

placement of the dish in the upright position.

• Using this method In situ comparison can be done

with the standard drug easily. While it is not so in

the Agar ditch and the paper disc method.

4.5 FACTORS AFFECTING ZONE OF INHIBITION :

Following factors are important for the screening of the

chemical substances (looking promising as the potential

therapeutic agents). These variables are to be kept constant

for the authentic and reproducible results.

4.5.1 Ingredients of culture media :

Many substances are present in culture media, which

may affect the zone of inhibition. Common ingredients such

as peptone, agar, etc. may vary in their contents. Many of

these minerals may influence the activity of some

antimicrobials. It is well known that Ca, Mg, Fe, etc. ions

affect the sensitivity of zone produced by the tetracycline,

gentamycin. NaCl reduce the activity of amino glycosides

and enhances the effect of fungicides.

4.5.2 Choice of media :

Consistent and reproducible results are obtained in

media prepared especially for sensitivity testing. The plates

must be poured flat with an even depth.

4.5.3 Effect of pH :

The activity of amino glycosides is enhanced in

alkaline media and reduced in acidic media. Exactly the

reverse is found true for the drugs of tetracycline family.

4.5.4 Size of inoculums :

Although large number of organisms does not markedly

affect many antibiotics, all inhibition zones are diminished

by heavy inoculum. The ideal inoculum is one, which gives

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an even dense growth without being confluent. Overnight

broth cultures of organisms and suitable suspensions from

solid media can be diluted accurately to give optimum

inoculum for sensitivity testing.

4.6 BACTERIA :

Bacteria are microscopic living creature classified in

the prokaryotic group of organisms. Many bacteria are

useful in our daily life from making the food like curd to the

production of pharmacological compound like antibiotics

and enzymes. On other side, many bacteria are found to be

pathogenic to the human and animal.

They cause the illness in human beings through the

infection. For treatment of the disease many natural and

synthetic compounds are currently used depending upon

the types of bacteria involved in the disease.

Classification of bacteria are based on several

characteristics exhibited by bacteria. However, the

frequently used is the Gram reaction. It was developed by

the Danish physician Christian Gram in 1884, discovered a

stain known as Gram stain, which can divide all bacteria

into two class “Gram positive” and “Gram negative”. The

Gram-positive bacteria resist discoloration with acetone,

alcohol and remain stained (methyl violet) as dark blue

colored, while Gram-negative bacteria are decolorized and

stains pink with the counter stain safranine. Bacteria are

also classified on the basis of their potency to cause the

disease. Those which are responsible for creating the

disease are known as a pathogenic and rests of other are

known as non-pathogenic

4.6.1 Pathogenic bacteria :

Those bacteria which cause the disease in the human

called the pathogenic bacteria. Many groups of bacteria are

Chapter -4


responsible to cause the disease in human, some are

obligatory pathogenic, while some are opportunistic

pathogenic.

4.6.2 Gram Negative Bacteria :

Pseudomonas aeruginosa is Gram negative opportunistic

pathogenic which is able to cause infection when the

natural resistance of the body is low. They are mostly

associated with hospital infection (nosocomial) and post

burn infection. They also cause infection of middle ear, eye

and urinary tract and also reported to cause the diarrhea

and pneumonia.

Escherichia coli is one of the most studied and well

characterized bacteria and opportunistic pathogenic cause

the traveler diarrhea and urinary tract infection. E.coli is

Gram negative, short rod, non-capsulated and non-

sporulated bacteria.

Proteus vulgaris is associated with urinary tract infection

as well as infection of wounds and burns. It is also

responsible for summer diarrhea and infantile diarrhea.

They are short rod, highly motile, Gram negative bacteria.

Organisms are non-sporulated and non-capsulated.

4.6.3 Gram positive bacteria :

Bacillus megaterium : It is one of the representative

species of the wide studied genus Baccillus bacteria and are

rod shape with round shape ends. Cells are non capsulated

but are motile and produce the spore. They are non

pathogenic but some strains cause the infection.

Staphylococcus aureus are Gram positive coccoi having a

round or oval shape with size of 0.8 to 0.10 µm in diameter.

They are non motile, non-sporulated and most of the strains

are non-capsulated but few strains show the capsule. They

are responsible for the pyogenic infection of the skin and

Chapter -4


also cause the infection like staphylococcal pneumonia,

wound infection, post burning and post surgical infections.

Colonies are smooth, raised, glistening, circular, entire

and translucent, and single colony may obtain a size of 6-8

mm in diameter on non selective media used for the

propagation of Staphylococci.

Facultative anaerobes : Growth is best under aerobic

condition. Growth in the anaerobic portion of a semisolid

thioglycolate medium is rapid and uniformly dense.

Terminal pH in glucose broth under anaerobic condition, is

4.3- 4.6. Catalase is produced by cell growing aerobically. It

is immunologically distinct from the catalase of other

species, as determined by double immuno diffusion and

micro complement fixation analyses. Catalase may be

absent in respiratory deficient mutants.

At least four different hemolysins (exotoxins) are

produced, including -, β-, - and δ- hemolysins. Nearly all

strains produce one or a combination of several hemolysins.

β-Hemolysin is produced more frequently by strains of

animal origin; it is a phospholipase C, specific for

sphingomyelin, and may be referred to as sphingomyelinase

C.

Most strains are susceptible to novobiocin (MIC < 0.4

μg/ml). Novobiocin resistance when it occurs is usually

chromosomally determined. Susceptibility to benzylpencillin

(penicillin G), erythromycin, tetracycline, chloremphenicol

and streptomycin is variable. Resistance to one or more of

these antibiotics and/or heavy metals (e.g. mercury, lead,

cadmium, arsenite, bismuth, antimony) is often plasmid

determined. Resistance to penicillin is due to production of

β-lactamases (penicillinase).

On the basis of phenotypic characterization,

Staphylococcus aureus strains living on certain different

Chapter -4


host species (e.g. cattle, pigs, hares, poultry, humans) can

be distinguished from one another and may be regarded as

representing different ecovars (or biovars). The distinction

of ecovar in is made on the basis of susceptibility to phase,

antigenic components of the cell wall, nutritional

requirement, coagulation of different plasmas, hemolysins,

fibronolysin activity, serological differences of nucleases

and crystal violet type. Cross contamination may give rise to

the isolation of more then one ecovar from an individual

host species (e.g. different species of animals on the same

farm, humans and their pets or farm animals).

4.7 ANTIMICROBIAL ESSAY :

The antimicrobial assay [12] is essential to evaluate

the compound against the microorganisms, the entire

procedure furnished in brief as under.

1. Method employed : Agar Cup Technique.

2. Concentration of the compound : 100 ppm.

3. Culture used : Gram +Ve and Gram –Ve bacteria.

4. Control : DMSO.

6. Results of inhibitory activity : Diameter of the inhibition

of zone measured in mm.

4.8 ORIGIN OF METHICILLIN RESISTANT STAPHYLO

COCCUS AUREUS (MRSA) :

As the new antibiotics introduced Staphylococcus

aureus has evolved resistance against those antibiotics. In

the late 1930’s, sulphonamides offered the first challenge to

S. aureus, but they failed because of their poor clinical

performance in the presence of pus, and the acquisition of

resistance by Steph bacteria [13].

As the progressive development of antibiotic, the

introduction of benzylpenicillin (Penicillin G) in to clinical

Chapter -4


practice in the early 1940s, dramatically improved this

prognosis. However, in 1942 resistance to this compound

was described, in a few S. aureus isolates. The mechanism

of resistance in Staphylococci was clarified shortly after

kirby recognized that resistant S. aureus produced a

penicillin inactivator. Bondi and Dietz identified the

inactivator to be penicillinase, an enzyme. 80% of S. aureus

showed resistance to benzyl penicillin by Melinda et al .

Along with it S. aureus had also acquired resistance to most

of the available antibiotics [14].

The isolation of the penicillin precursor 6-amino

penicillanic acid in 1959 made production of semisynthetic

penicillins possible. Modification of acyl side chain resulted

in steric protection of the β-lactam ring which prevented

hydrolysis of β-lactamase. Methicillin and oxacillin were

first to be introduced for clinical use in the early 1960’s.

These new compounds and related congeners like nafcillin,

cloxacillin, dicloxacillin provided a temporary solution to

the problem of penicillin resistance in Staphylococci.

Resistance to them was recognized almost immediately [14].

4.9 MATERIALS AND METHOD :

4.9.1 Selection of compounds for the antimicrobial

assay :

The various synthesized sulfa hydrazone substituted

4-(3H)-quinazolinone heterocyclic derivatives were screened

with the help of virtual screening as described in the

Chapter-3 in this thesis. Thus, the carcinogenic and

mutagenic compounds were removed. Then the screened

twenty compounds were used for the present study as

shown in Table 4.1.

Chapter -4


Table 4.1 Screened compounds for antibacterial activity:

COMPOUND ID

IUPAC NAME OF THE COMPOUNDS

Q1 4-amino-N-(2-methyl-4-oxoquinazolin-3-(4H)-yl)benzenesulfonamide.

Q2 4-amino-N-(6-bromo-2-methyl-4-oxoquinazolin-3-(4H)-yl)benzenesulfonamide.

Q8 4-amino-N-[2-(4-chlorophenyl)-4-oxoquinazolin-3-(4H)-yl]benzenesulfonamide.

Q9 4-amino-N-[6-bromo-2-(4-chlorophenyl)-4-oxoquinazolin-3-(4H)-yl]benzenesulfonamide.

Q10 4-amino-N-[6,8-dibromo-2-(4-chlorophenyl)-4-oxoquinazolin-3-(4H)-yl]benzenesulfonamide.

Q11 Ethyl-2-[(4-{[(2-methyl-4-oxoquinazolin-3-(4H)-yl)amino]sulfonyl}phenyl)hydrazono]-3-oxobutanoate.

Q12 2-{[4-(6-Bromo-2-methyl-4-oxo-4H-quinazolin-3-yl-sulfamoyl)phenyl]hydrazono}-3-oxobutyricacid ethylester.

Q22 4-[N'-(1-Acetyl-2-oxopropylidene)hydrazino]-N-(6-bromo-2-methyl-4-oxo-4H-quinazolin-3-yl)benzenesulfonamide.

Q25 4-[N'-(1-Acetyl-2-oxopropylidene)hydrazino]-N-(4-oxo-2-phenyl-4H-quinazolin-3-yl)benzenesulfonamide.

Q28 4-[N'-(1-Acetyl-2-oxo-propylidene)hydrazino]-N-[6-bromo-2-(4-chlorophenyl)-4-oxo-4H-quinazolin-3-yl]benzenesulfonamide.

Q31 N-(2-methyl-4-oxoquinazolin-3-(4H)-yl)-4-[(2E)-2-(4-oxo-2-thioxo-1,3-thiazolidin-5-ylidene)hydrazino]benzenesulfonamide.

Q35 N-(4-Oxo-2-phenyl-4H-quinazolin-3-yl)-4-{N'-[4- oxo-2-thioxothiazolidin-(5E)-ylidene]hydrazino}benzenesulfonamide.

Q41 4-{N'-[3-Methyl-5-oxo-1,5-dihydropyrazol-(4Z)-ylidene]hydrazino}-N-(2-methyl-4-oxo-4H-quinazolin-3-yl)benzenesulfonamide.

Q42 N-(6-Bromo-2-methyl-4-oxo-4H-quinazolin-3-yl)-4-{N'-[3-methyl-5-oxo-1,5-dihydropyrazol-(4Z)-ylidene]hydrazino}benzenesulfonamide.

Q45 4-{N'-[3-Methyl-5-oxo-1,5-dihydropyrazol-(4Z)-ylidene]hydrazino}-N-(4-oxo-2-phenyl-4H-quinazolin-3-yl)benzenesulfonamide.

Q46 N-(6-Bromo-4-oxo-2-phenyl-4H-quinazolin-3-yl)-4-{N'-[3-methyl-5-oxo-1,5-dihydropyrazol-(4Z)-ylidene]hydrazino}benzenesulfonamide.

Q61 4-[N'-(4,6-Dimethyl-2-oxo-2H-pyrimidin-5-ylidene)hydrazino]-N-(2-methyl-4-oxo-4H-quinazolin-3-yl)benzenesulfonamide.

Chapter -4


Q62 N-(6-Bromo-2-methyl-4-oxo-4H-quinazolin-3-yl)-4-[N'-(4,6-dimethyl-2-oxo-2H-pyrimidin-5-ylidene)hydrazino]benzenesulfonamide.

Q65 4-[N'-(4,6-Dimethyl-2-oxo-2H-pyrimidin-5-ylidene)hydrazino]-N-(4-oxo-2-phenyl-4H-quinazolin-3-yl)benzenesulfonamide.

Q68 N-[2-(4-Chlorophenyl)-4-oxo-4H-quinazolin-3-yl]-4-[N'-(4,6-dimethyl-2-oxo-2H-pyrimidin-5-ylidene)hydrazino]benzenesulfonamide.

4.9.2 Selection of bacteria :

The antibacterial activities of the synthesized

compounds were assayed against both the groups of

bacteria i.e. Grams positive and Grams negative to widen

the experiment approach.

4.9.3 Active culture preparation :

Nutrient broth was prepared by dissolving peptone

(0.5%), yeast extract (0.15%), beef extract (0.15%) and

sodium chloride (0.5%) in 100 ml distilled water. The pH of

the solution was adjusted to 7.2 by adding sodium

hydroxide solution (4%) and the resulting medium was

autoclaved for 15 min. at 15 psi. One day prior to the

experiment, all the cultures of bacteria were inoculated in

nutrient broth (inoculation medium) and incubated

overnight at 37 0C.

Similarly, nutrient agar medium was prepared with the

addition of 1.5% agar-agar powder as a solidifying agent in

to nutrient broth. Agar-agar was dissolved by heating the

medium followed by autoclave for 15 min. at 15 psi.

4.9.4 Antibacterial activity :

Each test compounds (1 mg) were dissolved in 1ml

DMSO, and 0.1ml from this prepared solution (100 μg) was

used for the testing of antibacterial activity. 24 hrs. old

culture was used as an inoculum and added aseptically to

the melted nutrient agar medium and mixed thoroughly to

get the uniform distribution of bacteria. Then medium was

Chapter -4


poured (25 ml in each dish) into Petri dishes and then

allowed to solidify at room temperature.

4.9.5 Cup borer method (Drug diffusion method) :

Solidified Plates were punched with a sterile cork borer

and scooping out the punched medium part made the cups.

The diameter of each cup was 6 mm. 0.1 ml of test

compound was added in to cups and DMSO was used as the

solvent control. The test samples were tested at a

concentration of 100 μg. The plates were allowed to keep at

low temperature for an hour in order to facilitate the

diffusion of the loaded solution. Then the plates were

incubated at 37 0C for 48 hrs.

4.10 RESULTS AND DISCUSSION :

Bacillus megaterium and multi-drug resistance

Staphylococcus aureus (MRSA) strain were used as a Gram

positive whereas, Escherichia coli, Pseudomonas aerugenosa

and Proteus vulgaris were included as a Gram negative. In

the present study MRSA was previously isolated form the

medical waste of two different hospitals and tested against

commercially available 12 different antibiotics, result of the

test indicates resistance against all the tested antibiotics

[16].

Overnight growth of all the bacteria was found enough

to use 0.1 ml inoculum in the assay medium. All the

bacteria were indicated the more or less equal growth.

Nutrient agar plates were indicated the lawn grown.

4.10.1 Measurement of antibacterial activity :

The zone of growth inhibition was observed and

measured in millimeters (mm) using antibiotic zone reader.

Zone of DMSO was considered as a control and subtracts

that with the zone of compounds. Zone against different

bacteria were compared to get the better ideas regarding the

Chapter -4


antibacterial activity of specific compounds with particular

bacteria as shown in Table 4.2.

Table 4.2 Zone of growth inhibition in mm :

Comp.

ID

Gram Negative Gram Positive

E.coli

(mm)

P.vulgaris

(mm)

P.

aerug

enosa

(mm)

B.mega

terium

(mm)

MRSA

Starain

1(mm)

MRSA

Starain

2(mm)

Q1 7 7 7 8 6 4

Q2 7 8 8 9 8 0

Q8 12 8 9 0 0 0

Q9 9 11 1 7 7 0

Q10 8 18 21 8 9 8

Q11 11 9 12 7 7 1

Q12 10 7 11 6 8 0

Q22 7 6 7 8 16 0

Q25 7 5 7 0 0 18

Q28 9 9 10 7 10 1

Q31 11 7 5 0 0 0

Q35 1 6 5 8 1 0

Q41 11 10 11 8 9 0

Q42 7 10 6 10 0 0

Q45 2 5 1 3 0 0

Q46 3 0 2 1 0 0

Q61 7 8 6 7 2 0

Q62 1 2 0 3 0 0

Q65 2 0 3 1 1 0

Q68 0 2 1 4 0 0

Chapter -4


4.10.2 Comparative analysis :

The normal strain of E. coli was also similarly inhibited

by all the compounds included in study within the range of

0 to 12 mm. Compound Q8 was found to be best inhibitor

with zone diameter of 12 mm followed by compound Q11,

Q31 and Q41. The least inhibition was found for Q62 and

Q68 compounds as shown in Figure 4.1.

Figure 4.1 Zone of inhibition of E. Coli.

Antimicrobial inhibition study with P. Proteus

demonstrated the activity in the range of 0 to 18 mm. The

compound Q10 was found to have the maximum

antimicrobial potential with zone inhibition of 18 mm,

followed by Q41, Q42 and Q11. The lowest inhibition was

reported for the compound Q46 and Q65 as shown in Figure

4.2.

Chapter -4


Figure 4.2 Zone of inhibition of P. vugaris

The entire compounds were found to inhibit growth of

Pseudomonas aerugenosa except compound Q8 in the range

of 0 to 21 mm. Maximum inhibition with zone diameter of

21 mm was recorded of compound Q10, followed by Q11,

Q12, Q28 and Q41 while minimum was for compounds Q45,

Q62 and Q68 as shown in Figure 4.3.

Figure 4.3 Zone of inhibition of P. aerugenosa

Escherichia coli, P. Proteus and Pseudomonas aerugenosa

was good inhibited by the compound Q8, indicate that

Chapter -4


compound Q8 was active against Grams negative bacteria

but not active against Gram positive bacteria like Bacillus

megaterium and MRSA strains.

In Bacillus megaterium, highest growth was found in

compound Q42 (Figure 4.4).

Figure 4.4 Zone of inhibition of B.megaterium

Antimicrobial activities of all the synthetic compounds

were found different against tested organisms. The above

study was also supported by Habib N. S. et al. [15].

Compounds Q1, Q2, Q9, Q11, Q12, Q22, Q28, and Q41 were

active against only one MRSA strain 1. Where as compound

Q1 and Q10 inhibited all the bacteria including MRSA stains

2. Therefore, both the compounds were active against all the

tested organisms. Similarly, compound Q25 was also active

against most of the strains of bacteria, but it was not good

inhibitor for Bacillus megaterium and one MRSA strain 1 as

shown in Figures 4.5 & 4.6.

In contrast to wide range inhibition of all the

compounds against all normal isolates the wide variation

was recorded for two MRSA strain 1 and MRSA strain 2. The

above study was also supported by Malik N. et al. [16].

Chapter -4


Figure 4.5 Zone of inhibition of MRSA stains 1

Figure 4.6 Zone of inhibition of MRSA stains 2

Thus, the present synthetic compounds have a potency

to use as a drug against MRSA strains and compounds may

have a future prospect to develop against MRSA strains,

Chapter -4


which are often difficult to treat but these synthetic

compounds are active against the MRSA strains.

4.11 CONCLUSION :

Based on the above study, it is to be concluded that

the antibacterial activities of the compounds were

influenced by the halogen group i.e. microbial activity was

enhanced on substitution of the bromine group, whereas

declining in the activity was observed in case of chlorine

group substitution in the present study. Moreover, bromine

groups were active against most of the bacteria as well as

MRSA strains. So it proves that, complexity of the

compound does not increase the antibacterial activity but

the activeness depends on the types of the substituted

groups and their position in the structure.

Escherichia coli was maximum inhibited by compound

Q8. Proteus vulgaris and Pseudomonas aerugenosa was

maximum inhibited by the compound Q10, and Bacillus

megaterium, highest inhibited by compound Q42.

Escherichia coli, Proteus vulgaris and Pseudomonas

aerugenosa was good inhibited by the compound Q8,

indicate that compound Q8 was active against Grams

negative bacteria.

The different pattern of inhibition with two MRSA

strains isolates from the diverge location indicates the

biochemical heterogeneity and drug resistance development

among both the isolates. The result indicates the good

potentiality of the compound Q1 and Q10 as a potent

candidate for the development of novel antimicrobial against

multidrug resistant strains of MRSA. Even currently many

reports are available regarding the sensitivity of MRSA

against quinazolinone derivatives. Therefore, the work

incorporated in the present thesis also supports this

Chapter -4


concept of inhibition of MRSA by the quinazolinone

derivatives.

MRSA strains are not easy to inhibit them. Although

three synthesized compounds inhibited the growth of MRSA

strain, these compounds are Q25 followed by Q10 and Q1

suggest that it may have a future potency to use against the

MRSA infection as a magic bullet to treat them in case of

hospital acquired infection of MRSA. However, further study

needs to evaluate the compounds in details.

Chapter -4


4.12 REFERENCES :

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Chapter -4


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