Synthetic strategy for the synthesis of thiophene 2-carboxamide derivatives substituted with hydroxyl, methyl and amino groups at position-3 was proposed. The strategy includes the cyclization of the precursor ethyl 2-arylazo-3-mercapto-3-(phenylamino)acrylate derivatives, 2-acetyl-2-arylazo-thioacetanilide derivatives and N-aryl-2-cyano-3-mercapto-3-(phenylamino)acrylamide derivatives with N-(4-acetylphenyl)-2-chloroacetamide in alcoholic sodium ethoxide. IR, 1H NMR, and mass spectroscopic analyses were used to characterize the synthesized derivatives. In addition, molecular, electronic properties of the synthesized products were studied by the density functional theory (DFT) where they exhibited close HOMO–LUMO energy gap (ΔEH-L) in which the amino derivatives 7a-c have the highest while the methyl derivatives 5a-c were the lowest. Using the ABTS method, the antioxidant properties of the produced compounds were evaluated, where amino thiophene-2-carboxamide 7a exhibit significant inhibition activity 62.0% compared to ascorbic acid The antibacterial activity against two pathogenic Gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis) and two of pathogenic Gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa) revealed that 7b records the highest activity index compared to ampicillin 83.3, 82.6, 64.0, 86.9%, respectively. Furthermore, the thiophene-2-carboxamide derivatives were docked with five different proteins with the use molecular docking tools and the results explained interactions between amino acid residue of enzyme and compounds. Compounds 3b and 3c showed the highest binding score with 2AS1 protein. Graphical Abstract

4-Aminobenzohydrazide (1) undergoes chloroacetylation twice, at the primary amine and hydrazide-NH2 functional groups. The conforming bis-chloroacetamide derivative 3 was reacted with different sulfur reagents (namely, 2-mercaptobenzothiazole, 6-amino-2-mercaptopyrimidin-4-ol, and 2-mercapto-4,6-dimethyl-nicotinonitrile) to give new bis-sulfide compounds 5, 7 and 9, respectively. The newly synthesised bis-chloroacetamide and corresponding sulfides were screened for anti-microbial and antioxidant potential. The sulfide derivative 7 exhibited the most potent activity against Staphylococcus aureus and Pseudomonas aeruginosa. It shows inhibition activities of 83.4% and 78.8%, respectively. Moreover, the sulfide derivative 7 showed the highest antioxidant activity with an inhibition ratio of 85.9%, which is close to L-ascorbic acid.

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