β-Lapachone (B-LAP) is a natural naphtaquinone with established anti-oxidative stress and anti-cancer activities. We aimed to investigate B-LAP protective potential against doxorubicin (DOX)-induced nephrotoxicity in mice. The mice received an oral dose of B-LAP followed by a single intraperitoneal injection of 20 mg/kg DOX a day later. They were then treated for 4 days with 1.25 mg/kg, 2.5 mg/kg, and 5 mg/kg doses of B-LAP. Renal levels of NAD/NADH ratios, p-AMPKα, p-NF-κB p65, inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6) along with renal expressions of TNF-α, IL-1β, and IL-6 were examined. Serum levels of kidney function markers as well as renal histopathology were also investigated. In addition to increasing the activities of p-AMPKα, B-LAP elevated NAD/NADH ratios in the kidneys and decreased the renal levels of nuclear p-NF-κB and its correspondent downstream effectors TNF-α, IL-1β, IL-6, and iNOS in the kidneys. Also, B-LAP effectively ameliorated renal architectural changes and attenuated serum levels of urea, creatinine, and cystatin C. Collectively, these findings suggest the protective actions of B-LAP against DOX-induced nephrotoxicity in mice.