Chaihu-Shu-Gan-San (CSGS) is a famous classic traditional Chinese medicines (TCM) formula for treatment of liver stagnancy recorded in a famous book of traditional Chinese medicine, Jing Yue Quan Shu published in 1624. It has been extensively accepted as an antidepressant in China and its mechanism of action is still not clear. Previously we have found that hepatic injury happens in chronic unpredicted mild stress (CUMS). Thus, the protection of CSGS against hepatic injury induced by CUMS treatment was explored by metabonomics study and gene expression of the rat liver tissue. The results indicated that CSGS improved 8 of the 18 perturbed potential biomarkers in liver tissues of rats treated with CUMS, and involved in regulating phospholipids and bile acid metabolism against hepatic injury induced by CUMS in rat. The expressions of two apoptosis associated genes (Bcl-2 and Bax) and four genes (Pnpla6, Pla2g15, Baat and Gad1) related to the perturbed metabolic pathways were further investigated by quantitative real-time polymerase chain reaction (qRT-PCR). Both metabonomics and studies of genetic influences on metabolites demonstrated that CSGS inhibited hepatocyte apoptosis, and regulated phospholipids and bile acid metabolism against hepatic injury induced by CUMS in rat. Exploring the protection of CSGS against hepatic injury related to depression further clarify the relationship between CUMS-induced depression and hepatic injury, and also provide a novel insight to understand the underlying antidepressive mechanism of CSGS.