Synthesis, in vitro bio-evaluation, and molecular docking study of thiosemicarbazone-based isatin/bis-Schiff base hybrid analogues as effective cholinesterase inhibitors

•Synthesis of thiosemicarbazone-based isatin/bis-Schiff base hybrid analogues.•Identification of a new class of acetylcholinesterase and butyrylcholinesterase inhibitors.•Analogue 1 proved to be most active acetylcholinesterase and butyrylcholinesterase inhibitors.•Docking study was conducted to interpret the obtained results. Analogues of isatin/bis-Schiff bases based on thiocarbohydrazone (1–16) were synthesized, characterised using various methods like NMR and HR-ESI-MS, and then tested against the acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) enzymes. All analogues showed good inhibitory potentials ranging from 0.20 ± 0.05 to 7.40 ± 0.20 M (for AChE) and 0.50 ± 0.05 to 9.40 ± 0.10 M (for BuChE) as compared to the reference drug Donepezil (IC50 = 2.16 ± 0.12 & 4.5 ± 0.11 M respectively). Analogue 1 was shown to be the most effective inhibitor of the AChE and BuChE enzymes among the synthesised analogues. The reason for Analogue 1 may be due to the role played by oxygen and the nitro motif in H-bonding with the active sites of AChE and BuChE enzymes. A structure–activity relationship (SAR) was developed based on nature, position, number, and the electron-donating and -withdrawing effects of substitution(s) on phenyl rings. Molecular docking studies were used to describe the binding interactions of the most active inhibitors with the active sites of AChE and BuChE.