Serious of some novel bis benzimidazole derivatives were designed and synthesized by nucleophilic substitution reaction of 5-(un)substituted-2-chloromethyl-1H-benzimidazole and 5-(un)substituted-2-mercapto-1H-benzimidazole in the presence of sodium in methanol. Spectral methods of analysis (FT-IR, 1H-NMR, and 13C-NMR) were used to confirm the structures of the synthesized compounds. Antibacterial activity were evaluated for most of the target compounds against four strains of bacteria including ( E.coli, P. aeruginosa ) as gram-negative bacteria and ( B. subtilis, S. aureus ) as gram-positive bacteria, the tested compounds showed various activity against both gram-negative and gram-positive bacteria used.