This work reports the production of nanoplastics (NPs) from polypropylene (PP) free of the antioxidant Irgafos® 168 (IRG) and alkane oligomers (ALK). PP pellets were milled into a powder with particle sizes in the 100–500 μm range. Additives and oligomers were removed using dichloromethane, and the powder exposed to UV irradiation, followed by filtration through 1 μm filters. PP suspensions, free of antioxidant and oligomers, were reloaded with IRG and ALK to their original commercial concentrations. This approach allowed testing the aquatic toxicity of IRG at concentrations compromised by water solubility limits. Toxicity assays using the cladoceran Daphnia magna with 24–48 h immobilization of neonates as endpoint showed toxicity for NPs containing IRG, with EC20 (48 h) in the 1.8–3.5 mg/L range, that corresponded to IRG exposure <1.2 μg/L. Suspensions of PP containing ALK, but not IRG, exhibited low toxicity (EC20 > 20 mg/L). The results allowed estimating the toxicity of IRG with a EC50 value of 3.3 ± 1.1 μg/L. Assays with different proportions of IRG and its oxidized form showed no differences. This work demonstrated the aquatic toxicity of IRG, for which there were no previous data, and developed a method for testing the toxicity of non-polar additives without being limited by their solubility.