The study aims to investigate the metabolic process of the antidepressant component (BS-60E) from Paeoniae Radix Alba in rats, and to compare the differences in the disposition of its metabolites between normal and depressed states. In this study, a chronic unpredictable mild stress (CUMS) depression model was established in rats. Normal and depressed rats were orally administered the BS-60E aqueous solution, and serum and urine samples were collected. The prototype constituents and metabolites were identified using UPLC-Q-TOF/MS combined with UNIFI software, and multivariate statistical analysis was employed to compare the differential constituents between normal and depressed rats. Animal experimental procedures were approved by the Animal Ethics Committee of the Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences & Peking Union Medical College (approval No.: SLXD-20211105019). The results showed that 9 and 28 prototype constituents, as well as 33 and 60 metabolites, were detected in serum and urine samples, respectively. The main metabolic pathways included phase Ⅰ reactions such as hydroxylation and desaturation, and phase Ⅱ reactions including glucuronidation and sulfation. Compared with normal rats, the levels of monoterpene constituents such as paeoniflorin and 1-O-β-D-glucopyranyl paeoniflorone were decreased in both serum and urine of depressed rats, while the levels of polyphenolic constituents such as methyl gallate and ethyl gallate were decreased in serum but increased in urine. These results indicate that the depressed state significantly alters the metabolic disposition of the main constituents of BS-60E in rats. This is manifested as reduced absorption of monoterpene constituents, leading to decreased exposure of prototype constituents in serum and urine, suggesting that their pharmacological effects may primarily depend on metabolites and are excreted via the kidneys in metabolized forms. Meanwhile, the renal excretion of polyphenolic constituents is accelerated, resulting in decreased serum concentrations and increased urinary excretion, suggesting that their prototype constituents may directly participate in the pharmacological process and are excreted in urine both as prototypes and metabolites.