Inter-appointment medication of the root canals with appropriate intracanal medicaments has been advocated to improve root canal disinfection. Graphene oxide (GO) has shown promising antimicrobial activity against a wide range of microorganisms, besides the capability of carrying antibiotics. The current study aimed to compare the antibacterial activity of double antibiotic paste (DAP) and GO per se and in combination (GO-DAP) against Enterococcus faecalis (E. faecalis).

A total of 108 extracted human mandibular premolars were contaminated with three-week-old E. faecalis and subjected to a primary microbial assessment. The samples were categorized into 15 groups concerning the intracanal medicament (DAP, GO, GO-DAP, and control) and contact time (1, 7, and 14 days). Then, the root canals were medicated, incubated, and resubjected to a secondary antimicrobial evaluation. The colony-forming units (CFU) were counted to calculate the antimicrobial efficacy. The data were analyzed via the Kruskal–Wallis test (α = 0.05).

GO-DAP was the only medicament that completely eradicated E. faecalis in 1 day. The percentage reduction of CFU/ml in the GO-DAP and DAP groups was higher than that in the GO group at all allocated contact times. Furthermore, a significant decrease of the CFU/ml was seen in the GO and DAP groups after 7 and 14 days of being medicated (P < 0.05).

Since GO-DAP improved root canal disinfection, this novel material can be introduced as a promising intracanal medicament against E. faecalis even in the short run.