BACKGROUND AND OBJECTIVES: Jaftex is a new herbal mouthwash that consists of the aqueous extract of Jaft oak, thymus, and the aqueous extract of Satureja Bachtiarica. Its cytotoxicity remains unknown. METHODS: The sample size was determined as 180 specimens as 3 specimens for each of the 60 different combinations of 2 mouthwashes, 2 cell lines, 3 exposure times (24, 48, 72 h), and 5 mouthwash doses. Also, there were 36 positive and negative controls. Jaftex 1% and chlorhexidine 0.2% were sterilized and diluted 1:2, 1:4, 1:8, 1:16, and 1:32 of the original concentration. Each drug dilution was made available in triplicate. OCC and HuGu cells were exposed to these concentrations at 37 °C for 24 h (n = 60), 48 h (n = 60), and 72 h (n = 60). The optical densities (ODs) were measured for each of the 180 experimental specimens and 36 positive and negative specimens. Also the IC50 was calculated. The results were analyzed (α = 0.05). RESULTS: The viability was much higher in the case of Jaftex compared with CHX. The viability was higher in OCC compared to HuGu. All pairwise comparisons between each two dosages were significant (all P values ≤ 0.013) in a way that by reducing the dosage, the viability increased. CONCLUSIONS: The toxicity of Jaftex was significantly lower than that of chlorhexidine. Also, the toxicity of both mouthwashes against the HuGu cell line was higher than the OCC-18 cell line. During 24, 48, and 72 h, the mouthwash toxicity increased significantly.