Methylphenidate hydrochloride (MPH), a widely prescribed pediatric drug for attention deficit hyperactivity disorder, induced liver adenocarcinomas in B6C3F1 mice exposed to 500 ppm in feed for 2 years (Dunnick and Hailey (1995) [14]). In order to determine if the induction of liver tumors was by a mutagenic mode of action, groups of male Big Blue (BB) mice (B6C3F1 background) were fed diets containing 50-4000 ppm MPH for 4, 12, or 24 weeks. At sacrifice, the livers were removed and the cII mutant frequency (MF) and spectrum of cII mutations were determined. In addition, the frequencies of micronucleated reticulocytes (MN-RETs) and normochromatic erythrocytes (MN-NCEs) were measured in peripheral blood erythrocytes as was the Hprt MF in splenic lymphocytes. Food consumption and body weight gain/loss were recorded weekly for each animal. The levels of MPH and RA were determined immediately before sacrifice in the serum of mice fed MPH for 24 weeks. A significant loss in body weights (p <or= 0.01) was found in mice fed the 2000 and 4000 ppm doses of MPH; however, there was no significant difference in the food consumption by any of the MPH-treated groups. The average liver cII MF in control animals was 20 +/- 4.5 x 10(-6) where as in MPH-treated animals, the average cII MFs ranged between 20+/-2.5 and 32 +/- 6.7 x 10(-6). None of the cII MFs in livers from any of the MPH treatment was significantly higher than the concurrent controls at either 4, 12 or 24 weeks. Further, there was no significant increase in either the Hprt MF or the micronucleus frequency at any time point in the treated animals. These results suggest that MPH is not mutagenic in mice and that the induction of tumors as previously reported in the liver is probably through a nongenotoxic mode of action.