This study investigates the variation of toxicity during ozonation of 2-chlorophenol (2-CP), 3-chlorophenol (3-CP) and 4-chlorophenol (4-CP) in neutral condition. Acute toxicity of pure chlorophenols (CPs) and their ozonated intermediates was evaluated by Microtox assay. The results revealed that the intermediates of oxidized CPs induced new toxicity during the early stage of ozonation, and the ozonated 2-CP showed higher degree of toxicity increase than 3-CP and 4-CP. The maximum toxicity normally occurred before the maximum color intensity was monitored, while ozone dosage applied was within 1 mg of ozone per mg of initial CPs. This increasing toxicity was mainly contributed from ozonated intermediates. Formation of chlorocatechols, chloromuconic acids and hydroxylated/chlorinated dimeric compounds were detected in ozonation of CPs. These chlorinated by-products may cause greater toxicity than the parent chlorophenols. The required ozone dosage to detoxify the CPs solution into a complete non-toxic condition follows the order: 4-CP > 3-CP > 2-CP.