Non-melanoma skin cancer (NMSC) accounts for >1 million new cases each year in the US alone suggesting that more approaches are needed for its prevention and control. Earlier studies by us have shown that silymarin (a crude form of biologically active silibinin with some other isomers), isolated from milk thistle, affords strong protection against ultraviolet (UV) radiation-induced NMSC in SKH-1 hairless mice; however, the molecular mechanisms of its efficacy are not known. Here, we assessed the effect of silibinin on UV-induced DNA damage and p53-p21/Cip1 accumulation, and their roles in UV-induced cell proliferation and apoptosis in SKH-1 hairless mouse epidermis. Topical application of silibinin prior to, or immediately after, UV irradiation resulted in a very strong protective effect against UV-induced thymine dimer positive cells in epidermis accounting for 76-85% (P < 0.001) inhibition. In other studies, silibinin treatment resulted in a further up-regulation of p53 by approximately 1.6-fold (P < 0.001) together with an increase ( approximately 2-fold, P < 0.001) in p21/Cip1 protein levels. Proliferative cell nuclear antigen staining showed that silibinin pre- or post-topical application significantly inhibits (40-52 and 20-40%, respectively, P < 0.001) UV-induced epidermal cell proliferation. In addition, silibinin strongly decreased UV-caused terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive apoptotic/sunburn cell formation (P < 0.001). These findings suggest that silibinin affords strong protection against UV-induced damage in epidermis by a decrease in thymine dimer positive cells and an up-regulation of p53-p21/Cip1 possibly leading to an inhibition in both cell proliferation and apoptosis. Comparable effects of silibinin following its pre- or post-UV application suggest that mechanisms other than sunscreen effect are operational in silibinin efficacy against UV-caused skin damages.