Objectives: The present study investigated the potential subacute toxicity of 1,4-dichlorobutane (1,4-DCB) by a 2-week repeated oral dose in male Sprague-Dawley rats. Materials and Methods: The test chemical was administered once daily by gavage to male rats at dose levels of 0, 74, 222, 667, and 2000 mg/kg/day for 2 weeks. All rats were sacrificed at the end of treatment period. During the test period, clinical signs, mortality, body weights, food and water consumption, urinalysis, hematology, serum biochemistry, gross findings, and organ weights were examined. Results: At 2000 mg/kg/day, treatment-related clinical signs, as evidenced by hypothermia, decreased locomotor activity, piloerection, lying on side, and prone position were observed. All the rats were found dead on test day 2. At 667 mg/kg/day, polyuria, suppressed body weight gain, food consumption, and spleen and thymus weights, and increased adrenal gland and liver weights were observed.Hematological and serum biochemical investigations revealed increases in the alanine aminotransferase, alkaline phosphataseand total bilirubinand decreases in the serum $Na^+$ level, white blood cell count and lymphocyte ratio. There were no treatment-related adverse effects in the 74 and 222 mg/kg/day groups. Conclusions: In the present experimental conditions, target organs were determined to be spleen, thymus,and liver. The no-observed-adverse-effect level was considered to be 222 mg/kg/day in male rats.