INTRODUCTION:

Microbial infections are one of the major reasons of the mortality and morality in the mankind. Microorganism like bacteria, virus and fungi are posing the significant problem to the healthcare systems. The discovery of the antibiotics and chemotherapeutic agents has significantly controlled this problem but due to continuous increase in the multidrug-resistant microbial strains infectivity of this microorganism is increased significantly. The resistance of these microbes towards antibiotics is one of the major reasons behind the increase in the infectivity. The irrational or continuous use of the agents like antibiotics and other chemotherapeutic agents leads to the strains which are totally resistance towards all the agents in the dosage regime which might leads to the critical situation. Development of the strains like methacilin resistant S. Aureus (MRSA), MDRTB, XDRTB, TDRTB pushing medicinal chemist for development of the new chemotherapeutic against all these resistant strains.

Development of the antibacterial agents is an interesting job which is mainly focused on the development of the agents which targets the replication of the bacteria. The replication cycle of the bacteria can be inhibited via inhibiting the critical enzymes which are important for the bacterial growth like DNA gyrase and Topoisomerase. DNA gyrase and topoisomerase are well known antimicrobial targets which are targeted by the one of the most widely utilized antimicrobial class fluroquinolones. Pyrimidine is one of most researched heterocyclic nucleus due to its wide spread presences in the nature and its importance in the overall development. Pyrimidine derivatives are often showed biological activities like antimicrobial, analgesic, anti-inflammatory, antidiabetic, antioxidant, antitumor, antibacterial. Keeping the biological significance in mind here we are reporting development of condensed pyrimidine derivatives using microwave assisted synthesis and their virtual and in vitro biological analysis as potent antimicrobial agents.

EXPERIMENTAL:

Design of condensed pyrimidine derivatives:

The optimized Condensed pyrimidine derivatives were designed on the basis of modification of the pyrimidine nucleus. The designed set of inhibitors has then scrutinized via computational analysis and the molecules which are shown promising results in the computational analysis are selected for the further analysis.

Docking analysis:²⁸

Molecular docking analysis was performed on the selected protein target Crystal Structure of E. coli 24kDa domain in complex with Clorobiocin (PDB ID 1KZN) was downloaded from the free protein databank www.rcsb.org. The docking analysis was performed using V life MDS. Biopredicta Module of the V life MDS was utilized for the docking analysis. The prepared protein structures were further utilized for docking simulation.

Synthesis of heterocyclic amine derivatives:

A mixture of thiosemicarbazide (0.01mol) and aryl carboxylic acid (0.01mol) was refluxed for 6–8 h using methanol: water (60:40) as solvent in the presence of conc. H₂SO₄ (10 drops). The mixture was then poured on crushed ice to get the amines.

Synthesis of condensed pyrimidine derivatives:

Equimolar quantities of the quinazoline derivatives and corresponding amines/heterocyclic amines are taken in to the round bottom flask and mixed well. 20ml of isopropyl alcohol is added into the reaction mixture and heated for 10 min in microwave. Reaction mixture is poured in to ice and separated product washed with water. All synthesized derivatives are shown in table no1.

Table no 1: Table showing synthesized lead derivatives

Molecule Code	R¹	R²
SAKB1
SAKB2
SAKB3
SAKB4
SAKB5	-H
SAKB6
SAKB7
SAKB8
SAKB9
SAKB10
SAKB11
SAKB12
SAKB13
SAKB14
SAKB15
SAKB16
SAKB17
SAKB18
SAKB19	-H
SAKB20	-H
SAKB21	-H
SAKB22	-H
SAKB23	-H
SAKB24	-H

N-(4-methylphenyl)-2-phenylquinazolin-4-amine (SAK18): Crystaline solid, point:.3499 (NH),334(C-H) 1H NMR (δ ppm) 2.48 (3H, s, CH₃), 7.2-7.5 (12 H m, ArH), 9.96 (1 H NH).

N-(4-bromo-phenyl)-2-phenylquinazolin-4-amine (SAKB10): Crystaline solid, point:.3400 (NH),2982(C-H) 1H NMR (δ ppm) 7.2-8.6 (13 H, m, ArH ), 8.61 (1 H NH )

N-(4-fluro-phenyl)-2-phenylquinazolin-4-amine (SAKB15): Crystaline solid, point:.3315.14 (NH),2900(C-H) 1H NMR (δ ppm) 7.2-8.2 (13H, m, ArH), 8.58 (1 H NH)

N-(4-chlorophenyl)quinazolin-4-amine (SAKB19): Crystaline solid, point:.3100 (NH),2800(C-H) 1H NMR (δ ppm) 6.9-8.4(13H, m, ArH ), 8.9(1 H NH)

Antimicrobial activity:

The inoculums of the microorganism were prepared from the bacterial cultures. 15ml of nutrient agar (Hi media) medium was poured in clean sterilized Petri plates and allowed to cool and solidify. 100 µl of broth of bacterial strain was pipette out and spread over the medium evenly with a spreading rod till it dried properly. Wells of 6mm in diameter were bored using a sterile cork borer. Solutions of all the extracts (1, 5 and 10mg/ml) in DMSO were prepared. 100µl of plant extracts solutions was added to the wells. The petri plates incubated at 370C for 24 h. streptomycin (1mg/ml) was prepared as a positive control DMSO was taken as negative control. Antibacterial activity was evaluated by measuring the diameters of the zone of inhibitions (ZI) all the determination were performed in triplicates.

RESULTS AND DISCUSSION:

Docking Analysis:

Grip based docking simulation are performed and all targeted molecules were found to be binding with selected target with good binding affinity and showing key interactions like hydrogen bond, Charge, Aromatic, hydrophobic and Vander wall interactions. All the molecules are having significant binding interaction with the targeted protein SAKB2 showed hydrogen bonding interaction with ARG76 and SAKB7 interacted via hydrogen bonding with ARG76, SAKB10 interacted with ARG76 with formation of hydrogen bond and, SAKB11 with ARG76, SAKB12 with ARG76 and ARG136 with formation of two hydrogen bond and, SAKB13 via formation of hydrogen bond with ASN46 anddARG76. Docking interactions are shown in figure no 1- 4.

Figure no 1: Docking Interactions of SAKB2

Figure no 2: Docking Interactions of SAKB7

Figure no 3: Docking Interactions of SAKB10

Figure no 4: Docking Interactions of SAKB11

Biological activity:

Anti-bacterial activity of the molecules was determined using disk plate method using Ciprofloxacin as standard. Anti-bacterial activity of the molecules was given in following table no 2 and figures no.5

Table No 2: Table showing antimicrobial activity of the synthesized derivatives.

Molecule Code	Zone of Inhibition
Molecule Code	B. Substilus	P. aeruginosa	E. coli	S. Aureus
SAKB1	03	04	05	02
SAKB2	14	15	12	11
SAKB3	02	02	01	01
SAKB4	05	06	04	03
SAKB5	04	05	06	04
SAKB6	04	01	01	02
SAKB7	13	10	14	12
SAKB8	17	16	14	12
SAKB9	11	10	13	16
SAKB10	14	12	14	15
SAKB11	12	14	11	15
SAKB12	14	12	14	15
SAKB13	15	14	15	16
SAKB14	04	05	06	07
SAKB15	14	12	11	10
SAKB16	04	05	07	08
SAKB17	11	10	14	12
SAKB18	04	05	01	03
SAKB19	12	14	15	16
SAKB20	15	20	09	17
SAKB21	14	15	14	12
SAKB22	03	02	01	02
SAKB23	05	02	04	03
SAKB24	01	02	02	04
Ciprofloxacin	26	21	28	27

Figure No 5: Figure Showing Antimicrobial Activity of synthesized derivatives.

CONCLUSION:

N,2-diphenylquinazolin-4-amine derivatives was synthesized using 4-chloro quinazoline derivatives and various aromatic amines. All the reaction was monitored using TLC and products are confirmed using analytical methods. Antibacterial activity of the all the derivatives was analysed using well diffusion method using Ciprofloxacin as a standard against B. Substilus, P.aeruginosa, E. coli and S. Aureus. All synthesized derivatives showed good activity against all the microorganisms. Among the compounds tested, SAKB2, SAKB7, SAKB8, SAKB9 SAKB10, SAKB11, SAKB12 and SAKB13 derivatives exhibited potent antimicrobial activity. The antibacterial activity revelled that the presence of the nitro, chloro, bromo substituted compounds are active than other derivatives. Our prediction is that by introducing these potent moieties into other new ring systems such as pyrazolopyrimidines and pyridopyrimidines may show even better antimicrobial activities.

ACKNOWLEDGEMENT:

Authors are thankful to Vlife Sciences for providing facility for research work.

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Received on 04.05.2021 Modified on 29.06.2021

Research J. Pharm. and Tech. 2022; 15(6):2422-2426.

DOI: 10.52711/0974-360X.2022.00403