Diethyl phthalate (DEP) has been widely used as a plasticiser in various consumer products, including cosmetics, personal care items, and pharmaceuticals, and recent studies reported a higher abundance of this priority phthalate in the aquatic environment. DEP is a potential endocrine disruptor, affecting immune systems in humans and wildlife even at low-level chronic exposure. As concern over phthalates increases globally, regulatory bodies focus more on their environmental impact. However, limited research is available, particularly using model organisms like planarians. Planarians are ideal for toxicological studies and may provide insightful information on pollutants' neurotoxic, developmental, and ecological effects, especially in freshwater environments where planarians play a vital role in ecosystem balance. Therefore, the objective of the current study was to examine the toxicity of DEP using the freshwater Dugesia sp., as an experimental animal. The LC50 for the test organism was calculated using DEP concentrations of 800, 400, 200, 100, and 50 µM, with an estimated LC50 of 357.24 µM. Furthermore, planarians were exposed to sub-lethal DEP concentration (178.62 µM) for one day as well as eight days to evaluate the impact of DEP on planarian locomotion, feeding behaviour, and regeneration ability. At sub-lethal concentration, locomotion and feeding ability were decreased, and regeneration was delayed. Furthermore, neuro-transmittance in planaria was altered by sub-lethal DEP concentration, as indicated by a reduced acetylcholinesterase (AChE) activity. DEP exposure induced oxidative damage in the tested planarians as shown by a marked increase in stress biomarkers, including lipid peroxidation levels and antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), peroxidase (POX), and glutathione S-transferase (GST). Our study revealed that DEP exposure may prove fatal to freshwater organisms, such as planarians. The observed alterations in behaviour and regeneration ability demonstrate the severity of the effects exerted by DEP as a toxicant in aquatic ecosystems, thereby indicating the need to restrict its usage to protect aquatic environments.