Background: Dihydrofolate reductase is one of the important enzymes for thymidylate
and purine synthesis in micro-organisms. A large number of drugs have been designed to inhibit
microbial DHFR but over the period of time, some drugs have developed resistance and cross
reactivity towards the enzyme. Over the past few decades, benzimidazoles, triazoles and their derivatives
have been grabbing the attention of the synthetic chemists for their wide gamut of antibacterial
and antifungal activities targeting microbial protein DHFR.
Objective: Our goal behind present investigation is to explore benzimidazoles class of drugs as microbial
DHFR inhibitors by studying ligand-receptor binding interactions, in vitro enzyme inhibition
assay and confirmation of anti-microbial activity against selected pathogenic microorganisms.
Methods: A library containing thirty novel 2,6-disubstituted 1H-benzimidazoles was synthesized
by one pot condensation of o-nitro aniline or 2,4-dinitro aniline with series of aldehydes or acetophenones
using Na2S2O4 or SnCl2 respectively and reflux for 5-6hr. Structures of compounds have
been confirmed by spectroscopic methods as 1H and 13C NMR, FT-IR and MS. In vitro DHFR inhibition
study was performed by using Epoch microplate reader and IC50 of the test compounds
was compared with Trimethoprim. In vitro antimicrobial activity was performed against selected
clinical pathogens by agar disk diffusion method and MIC (µg/mL) was reported.
Results: Moderate to good level of DHFR inhibition was observed with IC50 values in the range of
7-23 µM. Compounds B1, B19, B22, B24 and B30 expressed 1.1 to 1.4 folds more prominent
DHFR inhibitory activity as compared to standard Trimethoprim. Remarkable antimicrobial activity
was exhibited by B1, B19, B22, B24 and B30. Molecular docking study revealed perfect binding
of test ligands with key amino acids of DHFR as Phe31, Ile94, Ile5, Asp27, Gln32 and Phe36.
Conclusion: Nature of 1H-benzimidazole substituents at position 2 and 6 had influence over magnitude
and type of molecular binding and variation in the biological activity. The present series of
1H-benzimidazoles could be considered promising broad-spectrum antimicrobial candidates that
deserve in future for preclinical antimicrobial evaluation and development of newer antimicrobial
agents targeting microbial DHFR.