d-Methylphenidate (dexmethylphenidate; d-MPH) and its racemate d,l-methylphenidate (d,l-MPH) are currently prescribed for the chronic treatment of attention deficit hyperactivity disorder (ADHD) in children. Studies have shown that d-MPH is the pharmacologically active enantiomer for ADHD and is therefore the preferred drug for the treatment of ADHD symptoms. Although studies on the mutagenicity of d,l-MPH have been conducted, similar data for d-MPH are lacking. Therefore, d-MPH was evaluated in the bacterial reverse mutation and mouse lymphoma assays with and without S9 and in a bone marrow micronucleus test in male and female CD-1 mice. As a comparison, the l-enantiomer and racemate were also included in the assessments. While MPH-associated toxicity was observed in the mammalian tests, none of the three compounds tested induced mutagenic or clastogenic effects. Our present results along with published epidemiological data from patient populations are consistent with the conclusion that d-MPH and d,l-MPH do not present a carcinogenic risk to humans.