QXOH‐Levobupivacaine (LB) is a fixed‐dose combination of 35‐mM QXOH and 10‐mM LB. It was developed for perioperative analgesia because of its long‐acting analgesic effect. The purpose of this study was to evaluate the potential toxicity of QXOH‐LB in beagle dogs in accordance with the Guidance on the repeated‐dose toxicity published by the China Food and Drug Administration. Groups of five male and five female beagle dogs received normal saline, QXOH‐LB (2, 4, and 8 mg/kg, calculated as QXOH), QXOH (2, 4, and 8 mg/kg), or LB (2 mg/kg, equals the concentration of LB in 8‐mg/kg QXOH‐LB group) at the volume of 1 mL/kg once per day for 14 days through subcutaneous injection. No mortality was observed. Dogs in the control group as well as animals treated with 2‐mg/kg QXOH or QXOH‐LB exhibited normal behaviors. Clinical signs of toxicity in dogs treated with 4 and 8 mg/kg of QXOH or QXOH‐LB included decreased activity, unsteady gait, jerks, tremors, vocalization, emesis, ataxia, lateral/sternal recumbency, deep/rapid respiration, and gasping. Additionally, neurological function was found to be affected by QXOH and QXOH‐LB at the doses of 4 and 8 mg/kg. All clinical signs were recovered within 24 h. The no‐observed‐adverse‐effect level of QXOH and QXOH‐LB was considered to be 2 mg/kg. Toxicokinetic data showed that exposure to QXOH and LB increased as QXOH‐LB doses were increased from 4 to 8 mg/kg. There was no evidence of drug accumulation or any effect of gender.