This study aimed to compare the antimicrobial effect of an aqueous extract Red Roselle calyx (RE), Chlorhexidine (CH), Amoxicillin-clavulanic acid (ACA), Tetracycline (Tet), and Metronidazole (Met)on Streptococcus mutans (S. mutans), Staphylococcus aureus (S. aureus) and Enterococcus faecalis (E. faecalis) bacteria. The bacterial inhibition zones (BIZ)of the RE (25, 50, 75, 100) mg/ml and CH solutions (0.2%, 2%) were determined using the agar well diffusion method. Additionally, the susceptibility of the tested bacteria against (30 μg) of standard antibiotics of ACA, Tet, and Met was examined. The bacterial minimum inhibitory concentration (MIC) was measured using the Broth Micro dilution method (BMDM). All tests were carried out in triplicates, and water was considered the negative control. For S. mutans, the RE at 50 mg/ml or above concentrations displayed higher BIZ than 0.2% CH. 100 mg/ml of RE recorded a greater BIZ than the 2% CH. The greater BIZ against S. mutans was recorded by Tet. A comparable effect was found with 0.2% CH (75, 100) mg/ml of the RE against S. aureus. Greater BIZ for S. aureus and E. faecalis were reported for 100 mg/ml RE compared to the Tet and Met RE at 100 mg/ml inhibited the E. faecalis growth in a zone size comparable to the CH (0.2%, 2%).The RE with 50,100 mg/ml concentrations showed comparable antimicrobial effect to 0.2%, 2% concentrations of CH, respectively. As an herbal substitute for commercial disinfectants, the RE can be considered an effective final endodontic irrigant and dental mouthwash.