Fatty liver hemorrhagic syndrome (FLHS) is a metabolic disease mostly observed in laying hens that causes an economic toll on the poultry industry. To investigate the improving effects of biotin on FLHS in laying hens, a total of 135 Hy-Line Brown layers of 300-d-old were randomly divided into three groups and treated for 60 d. The hens from these three groups were fed with different diets: control group (the basal diet), pathology group [high-energy-low-protein diet (HELP)], and treatment group (HELP containing a biotin dosage of 0.3 mg kg^-1). The results showed that the mRNA expression level of apolipoprotein A I (apoA I) in pathology group significantly (P < 0.01) decreased on day 60 compared with the control group, while the mRNA level of apolipoprotein B₁₀₀ (apoB₁₀₀) increased significantly in pathology group on day 30, whereas it decreased significantly on day 60 (P < 0.05). Significantly increased mRNA levels of apoA I and apoB₁₀₀ were observed in treatment group compared with the pathology group on days 30 and 60 (P < 0.05 or P < 0.01). These results indicated that biotin could effectively alleviate the pathological changes and abnormal expression of apoA I and apoB₁₀₀ induced by FLHS, which might closely relate to the ability of biotin to promote egg production.