Polychlorinated biphenyls (PCBs) are ubiquitous and persistent environmental contaminants that disrupt endocrine function in biological systems, especially in the male reproductive system. Previous studies on the reproductive toxicity of PCBs have focused on the impairment of spermatogenesis, disruption of steroidogenesis, decreased sperm number, and infertility. Aroclor 1242 is a commercial mixture with an average of 42% chlorine by weight. The purpose of the present study was to elucidate the hazardous effects of Aroclor 1242 on Leydig cells through an evaluation of cell viability, lipid peroxidation, hydroxyl radicals, H2O2 production, antioxidant enzymes, and steroidogenic enzymes. Leydig cells were exposed to Aroclor 1242 for 24 h under basal and luteinizing hormone-stimulated conditions at different concentrations (ranging from 10-16 M to 10-6 M). After incubation, Leydig cells were measured for cell viability, lipid peroxidation, reactive oxygen species (hydroxyl radical and H2O2), antioxidant enzymes (catalase, superoxide dismutase, glutathione peroxidase, and glutathione-S-transferase), and steroidogenic enzymes (3β-hydroxysteroid dehydrogenase [HSD] and 17β-HSD). The results showed that cell viability was reduced only at Aroclor 1242 concentrations of 10-6 M and 10-8 M, whereas lipid peroxidation and reactive oxygen species increased relative to the concentration. Furthermore, antioxidant systems and steroidogenesis were interrupted to varying degrees, relative to the concentration. These findings suggest that exposure to Aroclor 1242 at high concentrations may result in detrimental effects to Leydig cell homeostasis. In addition, Aroclor 1242 may impair steroidogenesis, especially testosterone biosynthesis, by inhibiting two important steroidogenic enzymes.