Intratracheal instillation is a useful method for evaluating airway toxicity of various substances. However, there is limited information on this method in repeated dose toxicity studies. This study aimed to supplement existing background data by intratracheally administering distilled water for injection (DW) or phosphate-buffered saline (PBS) to four groups of six male SD rats each under anesthesia by inhalation of sevoflurane or isoflurane. Additionally, a non-intratracheal instillation group with inhalation anesthesia and a non-treatment control group with neither intratracheal instillation nor inhalation anesthesia were established. DW or PBS droplets were instilled via intratracheal intubation once a day, five days a week, for four weeks following inhalation anesthesia. The examination included hematology, blood chemistry, biochemical and cytological analysis of bronchoalveolar lavage fluid (BALF), organ weight measurement, gross necropsy, and histopathological examination of the lungs. No apparent abnormalities were observed in hematology, blood chemistry, or biochemical and cytological analysis of BALF. However, histopathological examination revealed perivascular/peribronchiolar eosinophil infiltration in the lungs induced by sevoflurane and isoflurane. The change was more pronounced with DW or PBS dosing, and was most severe in the DW groups, accompanied by focal inflammation. This study provides useful background data for conducting repeated dose toxicity studies via intratracheal instillation in rats.