Kaempferol is a flavonoid compound that has many functions, such as anti-inflammation and antioxidation. Acute liver failure (ALF) is a life-threatening illness accompanied by serious inflammation and extensive hepatocyte apoptosis. The aim of this study was to examine the therapeutic potential of kaempferol and its mechanism in ALF. In a murine ALF model induced by d-galactosamine (d-GalN, 700 mg/kg) / lipopolysaccharide (LPS, 10 μg/kg), mice were pretreated with kaempferol at 2 h before d-GalN/LPS administration and then sacrificed 6 h after d-GalN/LPS injection. Lethality, liver damage, endoplasmic reticulum(ER) stress, hepatocyte viability and apoptosis were evaluated. Whether pretreatment of kaempferol protected hepatocytes from ER stress-induced apoptosis was detected in vitro. Pretreatment of kaempferol decreased lethality, prolonged the survival time and significantly protected against liver injury, which was indicated by decreased transaminase levels and the well-preserved liver structure. The protective effect of kaempferol on the ALF mouse model was achieved by inhibiting hepatocyte apoptosis. Moreover, pretreatment of kaempferol increased the expression of glucose-regulated/binding immunoglobulin protein 78 (Grp78), decreased the expression of C/EBP-homologous protein (CHOP), and protected hepatocytes from ER stress-induced apoptosis in vitro. Our results showed that pretreatment of Grp78 siRNA partially negated the hepatic protection from kaempferol and reversed the inhibition of CHOP protein expression in d-GalN/LPS-induced ALF mice. In conclusion, kaempferol inhibits hepatocyte apoptosis to protect mice from liver failure by regulating the ER stress-Grp78-CHOP signaling pathway. Therefore, kaempferol may be used to treat ALF.