D-2-hydroxyglutaric (D-2-HGA) and L-2-hydroxyglutaric (L-2-HGA) acidurias are rare neurometabolic disorders biochemically characterized by increased levels of D-2-hydroxyglutaric acid (D-2-HG) and L-2-hydroxyglutaric acid (L-2-HG) respectively, in biological fluids and tissues. These diseases are caused by mutations in the specific enzymes involved in the metabolic pathways of these organic acids. In the present work, we first investigated whether D-2-HG and L-2-HGA could provoke DNA oxidative damage in blood leukocytes and whether L-carnitine (LC) could prevent the in vitro DNA damage induced by these organic acids. It was verified that 50 μM of D-2-HG and 30 μM of L-2-HG significantly induced DNA damage that was prevented by 30 and 150 μM of LC. We also evaluated oxidative stress parameters in urine of L-2-HGA patients and observed a significant increase of oxidized guanine species and di-tyrosine, biomarkers of oxidative DNA and protein damage, respectively. In contrast, no significant changes of urinary isoprostanes and reactive nitrogen species levels were observed in these patients. Taken together, our data indicate the involvement of oxidative damage, especially on DNA, in patients affected by these diseases and the protective effect of LC.