Objectives: The present randomised clinical study aimed to evaluate the effect of OneShape and ProTaper Next rotary systems on the removal of endotoxins from root canals of teeth with asymptomatic necrotic pulp. Materials and methods: 30 patients were randomly allocated into two groups (n = 15) according to the type of rotary system used in the mechanical preparation: group OneShape (OS) or group ProTaper Next (PN). Samples were collected before and after mechanical preparation and the quantification of endotoxins was conducted using the human enzyme-linked immunosorbent assay (ELISA) kit. Statistical analyses were performed using the Mann-Whitney U test to compare the two groups, and the Wilcoxon signed rank test to study the changes after treatment within each group. The significance level was set at P ≤ 0.05. Results: The group OS showed a statistically significant reduction in the endotoxin levels after treatment (P 0.001, effect size = 0.880). In the PN group there was a statistically significant reduction in endotoxin levels after treatment (P 0.001, effect size = 0.931). In the OS group a statistically significant median percentage reduction in endotoxin levels was observed compared with the group PN (P 0.05). Conclusions: ProTaper Next was more effective than OneShape in reducing endotoxins from root canals of teeth with asymptomatic necrotic pulp. Keywords: multi-file system, necrotic, OneShape, ProTaper Next, root canal, single-file system