Microwave-Assisted Organic Synthesis, structure–activity relationship, kinetics and molecular docking studies of non-cytotoxic benzamide derivatives as selective butyrylcholinesterase inhibitors

[Display omitted] •Microwave-Assisted Organic Synthesis is a facile method for the synthesis of benzamide derivatives.•Benzamides of 2,6-dichloroaniline showed selective in vitro butyrylcholinesterase (BCHE) inhibitory activity.•4-Bromosubstituted benzamide of 2,6-dichloroaniline (IC50 value = 0.8 ± 0.6 µM) was the most active analogue.•Enzyme kinetic studies of synthesized compounds showed mixed as well as uncompetitive types of inhibition. A series of benzamide derivatives 1–12 with various functional groups (–H, –Br, –F, –OCH3, –OC2H5, and –NO2) was synthesized using an economic, and facile Microwave-Assisted Organic Synthesis, and evaluated for acetylcholinesterase (ACHE) and butyrylcholinesterase (BCHE) activity in vitro. Structure–activity relationship showed that the substitution of –Br group influenced the inhibitory activity against BCHE enzyme. Synthesized compounds were found to be selective inhibitors of BCHE. In addition, all compounds 1–12 were found to be non-cytotoxic, as compared to the standard cycloheximide (IC50 = 0.8 ± 0.2 µM). Among them, compound 3 revealed the most potent BCHE inhibitory activity (IC50 = 0.8 ± 0.6 µM) when compared with the standard galantamine hydrobromide (IC50 = 40.83 ± 0.37 µM). Enzyme kinetic studies indicated that compounds 1, 3–4, and 7–8 showed a mixed mode of inhibition against BCHE, while compounds 2, 5–6 and 9 exhibited an uncompetitive pattern of inhibition. Molecular docking studies further highlighted the interaction of these inhibitors with catalytically important amino acid residues, such as Glu197, Hip438, Phe329, and many others.