Chungsimyeonja-tang (CSYJT) is an herbal decoction that consists with 12 herbal medicines. CSYJT has been shown to have anti-stress, anti-allergic and anti-oxidant effects. The aim of this study was to determine flavonoid contents in CSYJT and evaluate its protective effect against cisplatin induced nephrotoxicity using both in vitro (porcine renal epithelial cell; PK15 cell) and in vivo (Sprague Dawley rat) experiments. In the present study, thee mean contents of baicalin, wogonoside and baicalein in CSYJT were 14.65, 5.27 and 0.02 mg/g, respectively. The CSYJT extract treatment attenuated the following alteration in porcine renal epithelial (PK15) cell: the increase in reactive oxygen species (ROS), the glutathione depletion and the increase in p53 expression induced by cisplatin treatment. In the in vivo study, rats were orally treated with CSYJT extract once a day for 28 days. Five days before the last treatment, cisplatin (5 mg/kg) was intraperitoneally injected to induce acute renal failure. Increased blood urea nitrogen (BUN) and creatinine (CRE) levels after cisplatin treatment were ameliorated by pretreatment of CSYJT extract. In addition, lipid peroxidation was decreased and antioxidant enzyme (glutathione) was recovered in CSYJT pretreated kidney tissue. In histopathological examination, CSYJT pretreated group showed ameliorated pathological alteration after cisplatin injection with decreased apoptosis. Taken together, pretreatment of CSYJT could ameliorate cisplatin-induced nephrotoxicity.